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The present invention discloses a process for producing sodium 2-keto-L-gulonate from sorbitol and recovering 2-keto-L-gulonic acid in high recovery yields, by controlled cation exchange treatment of the micro-organism free fermentation broth and/or adjusting Ph of said purified fermentation broth, followed by direct crystallization of 2-keto-L-gulonic acid monohydrate. |
1. A process for producing sodium 2-keto-L-gulonate by fermentation and recovering 2-keto-L-gulonic acid monohydrate crystals, said process comprising: a) fermenting in a fermenting medium the presence of micro-organisms for converting sorbitol into at least 50 g/L sodium 2-keto-L-gulonate, b) removing the micro-organisms from the fermentation medium thereby obtaining a micro-organism-reduced fermentation broth, c) converting sodium 2-keto-L-gulonate into 2-keto-L-gulonic acid in the micro-organism-reduced fermentation broth, and removing proteins to a concentration below 2400 ppm (measured as nitrogen on dry substance) and obtaining a purified fermentation broth and/or adjusting the pH of the purified fermentation broth for avoiding the formation of vitamin C in a concentration higher than 3% (based on dry substance) during the subsequent evaporation of water, d) evaporating water from the purified fermentation broth to obtain a concentrated purified fermentation broth, and e) recovering 2-keto-L-gulonic acid monohydrate crystals by crystallization from the concentrated purified fermentation broth with a recovery yield for 2-keto-L-gulonic acid of 80% or higher. 2. A process according to claim 1, wherein in the step c) of said process the conversion into 2-keto-L-gulonic acid and the removal of proteins is obtained by ion exchange treatment consisting of cation exchange resin. 3. A process for producing sodium 2-keto-L-gulonate by fermentation and recovering 2-keto-L-gulonic acid monohydrate crystals, said process comprising: a) preparing a fermentation culture medium containing a nitrogen-source and, as a carbon source, sorbitol, b) innoculating the fermentation culture medium with micro-organisms for converting sorbitol into L-sorbose, c) allowing the micro-organisms to grow until at least 100 g/L L-sorbose is obtained in the fermentation medium, d) terminating the conversion of sorbitol into L-sorbose, e) innoculating the fermentation culture medium with micro-organisms for converting L-sorgose into sodium 2-keto-L-gulonate, f) allowing the micro-organisms to grow until at least 50 g/L sodium 2-keto-L-gulonate is obtained in the fermentation medium, g) removing the micro-organisms from the fermentation medium by filtration for obtaining a micro-organism-reduced fermentation broth, h) converting with cation exchange resin sodium 2-keto-L-gulonate into 2-keto-L-gulonic acid in the micro-organism-reduced fermentation broth, and removing proteins to a concentration below 2000 ppm (measured as nitrogen on dry substance) and obtaining a purified fermentation broth and/or adjusting the pH of the purified fermentation broth for avoiding the formation of vitamin C in a concentration higher than 2.5% (based on dry substance) during the subsequent evaporation of water, i) evaporating water from the purified fermentation and and obtaining a concentrated purified fermentation broth, and j) recovering 2-keto-L-gulonic acid monohydrate crystals by crystallization from the concentrated purified fermentation broth with a recovery yield fro 2-keto-L-gulonic acid of 80% and higher. 4. A process according to claim 3, wherein in step g) filtration is microfiltration, in step h) said purified fermentation broth comprises not more than 1800 ppm (measured as nitrogen on dry substance) proteins and adjusted pH is higher than 1.5, and in step j) recovery yield of 2-keto-L-gulonic acid is 85% or higher. 5. A process according to claims 3 or 4, wherein in step e) the micro-organism is a mixed culture of Gluconobacter oxydans, and Bacillus thuringiensis, and in said mixed culture the micro-organisms are present at the beginning of the growth in a ratio of Gluconobacter colonies to Bacillus colonies between 300:1 and 1:10. 6. A mixed culture of Gluconobacter oxydans, and Bacillus thuringiensis adapted for producing 2-keto-L-gulonic acid. 7. A mixed culture according to claim 6, wherein in said mixed culture the micro-organisims are present at the beginning of the growth in a ratio Gluconobacter colonies to Bacillus colonies of 25:1. 8. A Gluconobacter oxydans SCB 329 deposited at BCCM/′LMG on Apr. 24, 2001 under number LMG P-20356 for producing 2-keto-L-gulonic acid. 9. A Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20344 for producing 2-keto-L-gulonic acid. 10. A process for producing 2-keto-L-gulonic acid of micro-organism Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356, and micro-organism Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355. 11. A process according to claim 10, wherein said micro-organisms are applied in a mixed culture. 12. A process according to claim 1, wherein in step b) said fermentation broth is a micro-organism free fermentation broth. 13. A process according to claim 3, wherein in step b) said fermentation broth is a micro-organism free fermentation broth. 14. A process according to claim 3, wherein in step h), the concentration of vitamin C is not higher than 1% (based on dry substance). 15. A process according to claim 5, wherein said Gluconobacter oxydans comprises Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356, and said Bacillus thuringiensis comprises Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355. 16. A mixed culture of according to claim 6, wherein said Gluconobacter oxydans comprises Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356, and said Bacillus thuringiensis comprises Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355. |
<SOH> BACKGROUND OF THE INVENTION <EOH>It is known that 2-keto-L-gulonic acid can be produced by fermentation from L-sorbose and/or sorbitol. FR 1 376 741 relates to a process for the preparation of 2-keto-L-gulonic acid, which is an intermediate in the production of L-ascorbic acid (=vitamin C). It is demonstrated that the fermentative mixture can be used directly for esterification of 2-keto-L-gulonic acid into L-ascorbic acid. EP 0 518 136 describes a method for preparing 2-keto-L-gulonic acid that comprises cultivating a mixed culture of micro-organism (A) which belongs to the genus Gluconobacter or Acetobacter , and a micro-organism (B) capable of producing 2-keto-L-gulonic acid from L-sorbose and both of said micro-organisms are co-existing in the medium during at least part of the entire cultivation period. EP 0 278 447 relates to a fermentation process for producing 2-keto-L-gulonic acid by conversion of L-sorbose by means of mixed cultures of microorganisms comprising Gluconobacter oxydans and Bacillus megaterium. EP 0 972 843 relates to a continuous fermentation process for the manufacture of 2-keto-L-gulonic acid from sorbitol by fermentation with micro-organisms, in which process a nutrient medium containing sorbitol is incubated in a first fermentation vessel with a micro-organism capable of converting sorbitol to L-sorbose, and the resulting fermentation broth is transferred continuously to a second fermentation vessel where it is incubated with a micro-organism capable of converting L-sorbose to 2-keto-L-gulonic acid. EP 0 213 591 describes a process for producing 2-keto-L-gulonic acid and vitamin C respectively and the method further describes that the isolation of 2-keto-L-gulonic acid may be effected by the formation of a salt or by using the difference in properties between the product and impurities such as solubility, adsorbability and distribution coefficient between two solvents. Although the use of an adsorbent such as ion exchange resins is described as being one of the most convenient processes for isolation of the product, the 2-keto-L-gulonic acid thus obtained is in general not pure. EP 0 221 707 relates to a method for producing 2-keto-L-gulonic acid and describes a harvesting method wherein the culture broth is freed of cells by filtration, centrifugation or treatment with activated carbon and concentration of the solution. Solvent extraction, chromatography, precipitation or salting-out may be applied in a suitable combination and/or in repitition. WO 01/09152 relates to a process for the purification of 2-keto-L-gulonic acid by continuous liquid chromatography using a weakly basic ion exchange resin. It mainly relates to recovering 2-keto-L-gulonic acid substantially water-free from aqueous solutions. EP 0 359 645 relates to a process for obtaining pure 2-keto-L-gulonic acid from a fermentation broth containing the calcium salt of 2-keto-L-gulonic acid. Said process comprises separating the insolubles, concentrating the medium, precipitating calcium sulphate by adding concentrated sulphuric acid, treating with cation exchange resin, removing strong acid ions by anion exchanger, concentration and separating the 2-keto-L-gulonic acid by crystallisation. EP 0 805 210 relates to a process wherein sodium 2-keto-L-gulonate is crystallised from the fermentation broth, the obtained crystals are pulverised and suspended in a lower alcohol containing a water-free acid and finally the salt of 2-keto-L-gulonate is converted into a lower alkyl ester of 2-keto-L-gulonic acid. A further need exists to have a process for obtaining 2-keto-L-gulonic acid in high recovery yields and wherein the purification step is simple and easily reproducible. The current invention provides such a process. |
<SOH> SUMMARY OF THE INVENTION <EOH>The present invention discloses a process for producing sodium 2-keto-L-gulonate by fermentation and recovering 2-keto-L-gulonic acid monohydrate crystals and said process comprises the following steps: a) Fermentatively converting sorbitol into at least 50 g/L sodium 2-keto-L-gulonate, b) removing the micro-organisms from the fermentation medium thereby obtaining a micro-organism-reduced fermentation broth, preferably micro-organism-free fermentation broth, c) converting sodium 2-keto-L-gulonate into 2-keto-L-gulonic acid in the micro-organism-reduced fermentation broth, and removing proteins to a concentration below 2400 ppm (measured as nitrogen on dry substance) for obtaining a purified fermentation broth, and/or adjusting pH of purified fermentation broth for avoiding the formation of vitamin C in a concentration higher than 3% (based on dry substance) during the subsequent evaporation of water, d) evaporating water from the purified fermentation broth for obtaining a concentrated purified fermentation broth, and e) recovering 2-keto-L-gulonic acid monohydrate crystals by crystallisation from the concentrated purified fermentation broth with a recovery yield for 2-keto-L-gulonic acid of 80% or higher. The current invention further relates to a process wherein in said process the conversion into 2-keto-L-gulonic acid and the removal of proteins is effected by ion exchange treatment consisting of cation exchange resin. Furthermore, the current invention relates to a process comprising the following steps: a) Preparing a fermentation culture medium containing a nitrogen-source and as carbon-source sorbitol, b) Inoculating the fermentation culture medium with micro-organisms for converting sorbitol into L-sorbose, c) Allowing the micro-organisms to grow until at least 100 g/L L-sorbose is obtained in the fermentation medium, d) Terminating the conversion of sorbitol into L-sorbose, e) Inoculating the fermentation culture medium with a mixed culture of micro-organisms for converting L-sorbose into sodium 2-keto-L-gulonate, f) Allowing the micro-organisms to grow until at least 50 g/L sodium 2-keto-L-gulonate is obtained in the fermentation medium, g) removing the micro-organisms from the fermentation medium by filtration for obtaining a micro-organism-reduced fermentation broth, preferably micro organism-free fermentation broth, h) converting with cation exchange resin sodium 2-keto-L-gulonate into 2-keto-L-gulonic acid in the micro-organism reduced fermentation broth, and removing proteins to a concentration below 2000 ppm proteins (measured as nitrogen on dry substance) for obtaining a purified fermentation broth and/or adjusting pH of purified fermentation broth for avoiding during the subsequent evaporation of water the formation of vitamin C in a concentration higher than 2.5%, preferably not higher than 1% (based on dry substance), i) evaporating water from the purified fermentation broth for obtaining a concentrated purified fermentation broth, and j) recovering 2-keto-L-gulonic acid monohydrate crystals by crystallisation from the concentrated purified fermentation broth with a recovery yield for 2-keto-L-gulonic acid of 80% and higher. The current invention relates to a process wherein in step g) filtration is microfiltration, in step h) said purified fermentation broth comprises not more tin 1800 ppm (measured as nitrogen on dry substance) proteins and adjusted pH is higher than 1.5, and in step j) recovery yield of 2-keto-L-gulonic acid is 85% or higher. The current invention further relates to a process wherein in step b) the micro-organism is belonging to the genus Gluconobacter and in step e) the mixed culture of micro-organism is belonging to the genera Gluconobacter and Bacillus. The current invention specifically relates to a process wherein in step e) the micro-organism is a mixed culture of Gluconobacter oxydans , preferably Gluconobacter oxydans SCB 329 deposited under Budapest Treaty at BCCM/LMG (Belgian Coordinated Collections of Micro-organisms/Bacteria Collection Laboratorium voor Microbiologie Universiteit Gent by Cerestar Holding B.V. Nijverheidsstraat 1, NL-4551 LA Sas van Gent, The Netherlands) on Apr. 24, 2001 under number LMG P-20356, and Bacillus thuringiensis , preferably Bacillus thuringiensis (SCB 933) TCV 393 deposited under Budapest Treaty at BCCM/LMG (Belgian Coordinated Collections of Micro-organisms/Bacteria Collection Laboratorium voor Microbiologie Universiteit Gent by Cerestar Holding B.V. Nijverheidsstraat 1, NL-4551 LA Sas van Gent, The Netherlands) on Apr. 24, 2001 under number LMG P-20355, and in said mixed culture the micro-organisms are present at the beginning of the growth in a ratio Gluconobacter colonies to Bacillus colonies between 300:1 and 1:10. Furthermore, the current invention relates to a mixed culture of Gluconobacter oxydans , preferably Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356, and Bacillus thuringiensis , preferably Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355 for producing 2-keto-L-gulonic acid. Specifically the current invention relates to said mixed culture of micro-organisms wherein the micro-organisms are present at the beginning of the growth in a ratio of Gluconobacter colonies to Bacillus colonies of 25:1. The current invention relates to a Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356 for producing 2-keto-L-gulonic acid. The current invention further relates to a Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355 for producing 2-keto-L-gulonic acid. The current invention further relates to the use of micro-organism Gluconobacter oxydans SCB 329 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20356, and micro-organism Bacillus thuringiensis (SCB 933) TCV 393 deposited at BCCM/LMG on Apr. 24, 2001 under number LMG P-20355 for producing 2-keto-L-gulonic acid. Finally the current invention relates to the use wherein said micro-organisms are applied in a mixed culture for producing 2-keto-L-gulonic acid. detailed-description description="Detailed Description" end="lead"? |
Methods for diagnosing and treating neoplasias using nf-at transcriptions factors |
Described is a method for diagnosing a neoplasia by determining whether the level of one or more NF-AT transcription factor(s) is reduced. Moreover, methods for screening compounds which activate an NF-AT or which mimic the function of NF-AT are disclosed. Also disclosed are methods for the prevention or treatment of neoplasias by increasing the activity of an NF-AT. |
1. A diagnostic method comprising the step of determining in a sample taken from a patient the expression level and/or activity of one or more NF-AT transcription factor(s), and wherein a decrease or loss of the expression and/or activity of the NF-AT transcription factor(s) is indicative for the occurrence of a neoplasia. 2. The diagnostic method of claim 1, wherein the neoplasia is a neoplasia of hematopoietic cells, of chondrocytes, of osteocytes, of heart cells, muscle cells or blood vessel cells. 3. The diagnostic method of claim 1, wherein the neoplasia is a lymphoma. 4. The diagnostic method of claim 3, wherein the lymphoma is a T cell lymphoma a B-cell lymphoma, or Hodgkin lymphoma. 5. The diagnostic method of claim 1, wherein the sample is a biopsy. 6. The diagnostic method of claim 5, wherein the sample is a biopsy from a lymph node. 7. The diagnostic method of claim 1, wherein the sample is a blood sample. 8. The diagnostic method of claim 1, wherein the NF-AT transcription factor is selected from the group consisting of: (a) NF-ATc1; (b) NF-ATc2; (c) NF-ATc3; (d) NF-ATc4; and a combination thereof. 9. The diagnostic method of claim 1, wherein the decrease or loss of the expression and/or activity of the NF-AT transcription factor(s) is detected by determining the amount of RNA encoding the NF-AT transcription factor(s) in the sample. 10. The diagnostic method of claim 1, wherein the decrease or loss of the expression and/or activity of the NF-AT transcription factor(s) is detected by determining the amount of the NF-AT transcription factor(s) in the sample. 11. The diagnostic method of claim 1, wherein the decrease or loss of the expression and/or activity of the NF-AT transcription factor(s) is detected by determining the DNA binding or the transactivation activity of the NF-AT transcription factor(s) in the sample. 12. The diagnostic method of claim 1, wherein the decrease or loss of the expression and/or activity of the NF-AT transcription factor is determined by analysing the methylation state of the promoter of the NF-AT gene(s) or by detecting a loss of heterozygosity. 13. A diagnostic composition comprising a mixture of antibodies which are specific for more than one NF-AT transcription factors or a nucleic acid molecule specifically hybridizing with more than one nucleotide sequence encoding an NF-AT transcription factor or with a promoter sequence of more than one NF-AT gene. 14. A pharmaceutical composition comprising a polypeptide which comprises the C-terminal peptide of 246 amino acids of the isoform C of an NF-ATc1 transcription factor or a nucleic acid molecule encoding such a polypeptide and optionally a pharmaceutically acceptable carrier. 15. Use A method of preventing or treating neoplasia comprising administration of a pharmaceutically effective amount of the pharmaceutical composition of claim 14. |
Nucleic-acid associated proteins |
Various embodiments of the invention provide human nucleic acid-associated proteins (NAAP) and polynucleotides which identify and encode NAAP. Embodiments of the invention also provide expression vectors, host cells, antibodies, agonists, and antagonists. Other embodiments provide methods for diagnosing, treating, or preventing disorders associated with aberrant expression of NAAP. |
1. An isolated polypeptide selected from the group consisting of: a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-2, SEQ ID NO:4-13, SEQ ID NO:15-19, SEQ ID NO:21, SEQ ID NO:26, SEQ ID NO:28-29, and SEQ ID NO:31, c) a polypeptide comprising a naturally occurring amino acid sequence at least 93% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:23 and SEQ ID NO:25, d) a polypeptide comprising a naturally occurring amino acid sequence at least 95% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:3, SEQ ID NO:22, and SEQ ID NO:27, e) a polypeptide comprising a naturally occurring amino acid sequence at least 97% identical to the amino acid sequence of SEQ ID NO:30, f) a polypeptide comprising a naturally occurring amino acid sequence at least 99% identical to the amino acid sequence of SEQ ID NO:33, g) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and h) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. 2. An isolated polypeptide of claim 1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. 3. An isolated polynucleotide encoding a polypeptide of claim 1. 4. An isolated polynucleotide encoding a polypeptide of claim 2. 5. An isolated polynucleotide of claim 4 comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66. 6. A recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide of claim 3. 7. A cell transformed with a recombinant polynucleotide of claim 6. 8. (canceled) 9. A method of producing a polypeptide of claim 1, the method comprising: a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide, and said recombinant polynucleotide comprises a promoter sequence operably linked to a polynucleotide encoding the polypeptide of claim 1, and b) recovering the polypeptide so expressed. 10. A method of claim 9, wherein the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. 11. An isolated antibody which specifically binds to a polypeptide of claim 1. 12. An isolated polynucleotide selected from the group consisting of: a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-56 and SEQ ID NO:58-66, c) a polynucleotide complementary to a polynucleotide of a), d) a polynucleotide complementary to a polynucleotide of b), and e) an RNA equivalent of a)-d). 13. (canceled) 14. A method of detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 12, the method comprising: a) hybridizing the sample with a probe comprising at least 20 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide or fragments thereof, and b) detecting the presence or absence of said hybridization complex, and, optionally, if present, the amount thereof. 15. (canceled) 16. A method of detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 12, the method comprising: a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof, and, optionally, if present, the amount thereof. 17. A composition comprising a polypeptide of claim 1 and a pharmaceutically acceptable excipient. 18. A composition of claim 17, wherein the polypeptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. 19. (canceled) 20. A method of screening a compound for effectiveness as an agonist of a polypeptide of claim 1, the method comprising: a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting agonist activity in the sample. 21.-22. (canceled) 23. A method of screening a compound for effectiveness as an antagonist of a polypeptide of claim 1, the method comprising: a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting antagonist activity in the sample. 24.-25. (canceled) 26. A method of screening for a compound that specifically binds to the polypeptide of claim 1, the method comprising: a) combining the polypeptide of claim 1 with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide of claim 1 to the test compound, thereby identifying a compound that specifically binds to the polypeptide of claim 1. 27. (canceled) 28. A method of screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a sequence of claim 5, the method comprising: a) exposing a sample comprising the target polynucleotide to a compound, under conditions suitable for the expression of the target polynucleotide, b) detecting altered expression of the target polynucleotide, and c) comparing the expression of the target polynucleotide in the presence of varying amounts of the compound and in the absence of the compound. 29. A method of assessing toxicity of a test compound, the method comprising: a) treating a biological sample containing nucleic acids with the test compound, b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide of claim 12 under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide comprising a polynucleotide sequence of a polynucleotide of claim 12 or fragment thereof, c) quantifying the amount of hybridization complex, and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound. 30.-121. (canceled) |
<SOH> BACKGROUND OF THE INVENTION <EOH>Multicellular organisms are comprised of diverse cell types that differ dramatically both in structure and function. The identity of a cell is determined by its characteristic pattern of gene expression, and different cell types express overlapping but distinctive sets of genes throughout development. Spatial and temporal regulation of gene expression is critical for the control of cell proliferation, cell differentiation, apoptosis, and other processes that contribute to organismal development. Furthermore, gene expression is regulated in response to extracellular signals that mediate cell-cell communication and coordinate the activities of different cell types. Appropriate gene regulation also ensures that cells function efficiently by expressing only those genes whose functions are required at a given time. The cell nucleus contains all of the genetic information of the cell in the form of DNA, and the components and machinery necessary for replication of DNA and for transcription of DNA into RNA. (See Alberts, B. et al. (1994) Molecular Biology of the Cell, Garland Publishing Inc., New York N.Y., pp. 335-399.) DNA is organized into compact structures in the nucleus by interactions with various DNA-binding proteins such as histones and non-histone chromosomal proteins. DNA-specific nucleases, DNAses, partially degrade these compacted structures prior to DNA replication or transcription. DNA replication takes place with the aid of DNA helicases which unwind the double-stranded DNA helix, and DNA polymerases that duplicate the separated DNA strands. Transcription Factors Transcriptional regulatory proteins are essential for the control of gene expression. Some of these proteins function as transcription factors that initiate, activate, repress, or terminate gene transcription. Transcription factors generally bind to the promoter, enhancer, and upstream regulatory regions of a gene in a sequence-specific manner, although some factors bind regulatory elements within or downstream of a gene coding region. Transcription factors may bind to a specific region of DNA singly or as a complex with other accessory factors. (Reviewed in Lewin, B. (1990) Genes IV, Oxford University Press, New York, N.Y., and Cell Press, Cambridge, Mass., pp. 554-570.) The double helix structure and repeated sequences of DNA create topological and chemical features which can be recognized by transcription factors. These features are hydrogen bond donor and acceptor groups, hydrophobic patches, major and minor grooves, and regular, repeated stretches of sequence which induce distinct bends in the helix. Typically, transcription factors recognize specific DNA sequence motifs of about 20 nucleotides in length. Multiple, adjacent transcription factor-binding motifs may be required for gene regulation. Many transcription factors incorporate DNA-binding structural motifs which comprise either a helices or β sheets that bind to the major groove of DNA. Four well-characterized structural motifs are helix-turn-helix, zinc finger, leucine zipper, and helix-loop-helix. Proteins containing these motifs may act alone as monomers, or they may form homo- or heterodimers that interact with DNA. The helix-turn-helix motif consists of two α helices connected at a fixed angle by a short chain of amino acids. One of the helices binds to the major groove. Helix-turn-helix motifs are exemplified by the homeobox motif which is present in homeodomain proteins. These proteins are critical for specifying the anterior-posterior body axis during development and are conserved throughout the animal kingdom. The Antennapedia and Ultrabithorax proteins of Drosophila melanogaster are prototypical homeodomain proteins. (Pabo, C. O. and R. T. Sauer (1992) Annu. Rev. Biochem. 61:1053-1095.) Homeobox genes are a family of highly conserved regulatory genes that encode transcription factors. They are essential during embryonic development. They are important in limb formation and reproductive tract development. They function in uterine receptivity and implantation in mice and probably serve a similar role in humans (Daftary, G. S. and H. S. Taylor (2000) Semin. Reprod. Med. 18:311-320). Homeobox gene mutations play a role in susceptibility to autism (Ingram, J. L. et al. (2000) Teratology 62:393-405) and are implicated in human diseases, such as diabetes to cancer (Cillo, C. et al. (2001) J. Cell Physiol. 188:161-169). The helix-loop-helix motif (HLH) consists of a short a helix connected by a loop to a longer a helix. The loop is flexible and allows the two helices to fold back against each other and to bind to DNA. The protooncogene Myc, a transcription factor that activates genes required for cellular proliferation, contains a prototypical HLH motif. A zinc finger is a cysteine-rich, compactly folded protein motif in which specifically placed cysteines, and in some cases histidines, coordinate Zn +2 . Several types of zinc finger motifs have been identified. Though originally identified in DNA-binding proteins as regions that interact directly with DNA, zinc fingers occur in a variety of proteins that do not bind DNA (Lodish, H. et al. (1995) Molecular Cell Biology, Scientific American Books, New York, N.Y., pp. 447-451). For example, Galcheva-Gargova, Z. et al. ((1996) Science 272:1797-1802) have identified zinc finger proteins that interact with various cytokine receptors. The zinc finger motif, which binds zinc ions, generally contains tandem repeats of about 30 amino acids consisting of periodically spaced cysteine and histidine residues. Examples of this sequence pattern, designated C2H2 and C3HC4 (“RING” finger), have been described (Lewin, supra). Zinc finger proteins each contain an α helix and an antiparallel β sheet whose proximity and conformation are maintained by the zinc ion. Contact with DNA is made by the arginine preceding the α helix and by the second, third, and sixth residues of the α helix. Variants of the zinc finger motif include poorly defined cysteine-rich motifs which bind zinc or other metal ions. These motifs may not contain histidine residues and are generally nonrepetitive. The zinc finger motif may be repeated in a tandem array within a protein, such that the a helix of each zinc finger in the protein makes contact with the major groove of the DNA double helix. This repeated contact between the protein and the DNA produces a strong and specific DNA-protein interaction. The strength and specificity of the interaction can be regulated by the number of zinc finger motifs within the protein. Though originally identified in DNA-binding proteins as regions that interact directly with DNA, zinc fingers occur in a variety of proteins that do not bind DNA (Lodish, H. et al. (1995) Molecular Cell Biology, Scientific American Books, New York N.Y., pp. 447-451). For example, Galcheva-Gargova, Z. et al. (1996; Science 272:1797-1802) have identified zinc finger proteins that interact with various cytokine receptors. The C2H2-type zinc finger signature motif contains a 28 amino acid sequence, including 2 conserved Cys and 2 conserved His residues in a C-2-C-12-H-3-H type motif. The motif generally occurs in multiple tandem repeats. A cysteine-rich domain including the motif Asp-His-His-Cys (DHHC-CRD) has been identified as a distinct subgroup of zinc finger proteins. The DHHC-CRD region has been implicated in growth and development. One DHHC CRD mutant shows defective function of Ras, a small membrane-associated GTP-binding protein that regulates cell growth and differentiation, while other DHHC-CRD proteins probably function in pathways not involving Ras (Bartels, D. J. et al. (1999) Mol. Cell Biol. 19:6775-6787). Zinc-finger transcription factors are often accompanied by modular sequence motifs such as the Kruppel-associated box (KRAB) and the SCAN domain. For example, the hypoalphalipoproteinemia susceptibility gene ZNF202 encodes a SCAN box and a KRAB domain followed by eight C2H zinc-finger motifs (Honer, C. et al. (2001) Biochim. Biophys. Acta 1517:441-448). The SCAN domain is a highly conserved, leucine-rich motif of approximately 60 amino acids found at the amino-terminal end of zinc finger transcription factors. SCAN domains are most often linked to C2H2 zinc finger motifs through their carboxyl-terminal end. Biochemical binding studies have established the SCAN domain as a selective hetero- and homotypic oligomerization domain. SCAN domain-mediated protein complexes may function to modulate the biological function of transcription factors (Schumacher, C. et al. (2000) J. Biol. Chem. 275:17173-17179). The KRAB (Kruppel-associated box) domain is a conserved amino acid sequence spanning approximately 75 amino acids and is found in almost one-third of the 300 to 700 genes encoding C2H2 zinc fingers. The KRAB domain is found N-terminally with respect to the finger repeats. The KRAB domain is generally encoded by two exons; the KRAB-A region or box is encoded by one exon and the KRAB-B region or box is encoded by a second exon. The function of the KRAB domain is the repression of transcription. Transcription repression is accomplished by recruitment of either the KRAB-associated protein-1, a transcriptional corepressor, or the KRAB-A interacting protein. Proteins containing the KRAB domain are likely to play a regulatory role during development (Williams, A. J. et al. (1999) Mol. Cell Biol. 19:8526-8535). A subgroup of highly related human KRAB zinc finger proteins detectable in all human tissues is highly expressed in human T lymphoid cells (Bellefroid, E. J. et al. (1993) EMBO J. 12:1363-1374). The 2NF85 KRAB zinc finger gene, a member of the human ZNF91 family, is highly expressed in normal adult testis, in seminomas, and in the NT2/D1 teratocarcinoma cell line (Poncelet, D. A. et al. (1998) DNA Cell Biol. 17:931-943). The C4 motif is found in hormone-regulated proteins. The C4 motif generally includes only 2 repeats. A number of eukaryotic and viral proteins contain a conserved cysteine-rich domain of 40 to 60 residues (called C3HC4 zinc-finger or RING finger) that binds two atoms of zinc, and is probably involved in mediating protein-protein interactions. The 3D “cross-brace” structure of the zinc ligation system is unique to the RING domain. The spacing of the cysteines in such a domain is C-x(2)-C-x(9 to 39)C-x(1 to 3)-H-x(2 to 3)-C-x(2)-C-x(4 to 48)-C-x(2)-C. The PHD finger is a C4HC3 zinc-finger-like motif found in nuclear proteins thought to be involved in chromatin-mediated transcriptional regulation. GATA-type transcription factors contain one or two zinc finger domains which bind specifically to a region of DNA that contains the consecutive nucleotide sequence GATA. NMR studies indicate that the zinc finger comprises two irregular anti-parallel β sheets and an α helix, followed by a long loop to the C-terminal end of the finger (Ominchinski, J. G. (1993) Science 261:438-446). The helix and the loop connecting the two β-sheets contact the major groove of the DNA, while the C-terminal part, which determines the specificity of binding, wraps around into the minor groove. The LIM motif consists of about 60 amino acid residues and contains seven conserved cysteine residues and a histidine within a consensus sequence (Schmeichel, K. L. and M. C. Beckerle (1994) Cell 79:211-219). The LIM family includes transcription factors and cytoskeletal proteins which may be involved in development, differentiation, and cell growth. One example is actin-binding LIM protein, which may play roles in regulation of the cytoskeleton and cellular morphogenesis (Roof, D. J. et al. (1997) J. Cell Biol. 138:575-588). The N-terminal domain of actin-binding LIM protein has four double zinc finger motifs with the LIM consensus sequence. The C-terminal domain of actin-binding LIM protein shows sequence similarity to known actin-binding proteins such as dematin and villin. Actin-binding LIM protein binds to F-actin through its dematin-like C-terminal domain. The LIM domain may mediate protein-protein interactions with other LIM-binding proteins. Myeloid cell development is controlled by tissue-specific transcription factors. Myeloid zinc finger proteins (MZF) include MZF-1 and MZF-2. MZF-1 functions in regulation of the development of neutrophilic granulocytes. A murine homolog MZF-2 is expressed in myeloid cells, particularly in the cells committed to the neutrophilic lineage. MZF-2 is down-regulated by G-CSF and appears to have a unique function in neutrophil development (Murai, K. et al. (1997) Genes Cells 2:581-591). The leucine zipper motif comprises a stretch of amino acids rich in leucine which can form an amphipathic α helix. This structure provides the basis for dimerization of two leucine zipper proteins. The region adjacent to the leucine zipper is usually basic, and upon protein dimerization, is optimally positioned for binding to the major groove. Proteins containing such motifs are generally referred to as bZIP transcription factors. The leucine zipper motif is found in the proto-oncogenes Fos and Jun, which comprise the heterodimeric transcription factor AP1 involved in cell growth and the determination of cell lineage (Papavassiliou, A. G. (1995) N. Engl. J. Med. 332:45-47). The NF-kappa-B/Rel signature defines a family of eukaryotic transcription factors involved in oncogenesis, embryonic development, differentiation and immune response. Most transcription factors containing the Rel homology domain (RHD) bind as dimers to a consensus DNA sequence motif termed kappa-B. Members of the Rel family share a highly conserved 300 amino acid domain termed the Rel homology domain. The characteristic Rel C-terminal domain is involved in gene activation and cytoplasmic anchoring functions. Proteins known to contain the RHD domain include vertebrate nuclear factor NF-kappa-B, which is a heterodimer of a DNA-binding subunit and the transcription factor p65, mammalian transcription factor RelB, and vertebrate proto-oncogene c-rel, a protein associated with differentiation and lymphopoiesis (Kabrun, N. and P. J. Enrietto (1994) Semin. Cancer Biol. 5:103-112). A DNA binding motif termed ARID (AT-rich interactive domain) distinguishes an evolutionarily conserved family of proteins. The approximately 100-residue ARID sequence is present in a series of proteins strongly implicated in the regulation of cell growth, development, and tissue-specific gene expression. ARID proteins include Bright (a regulator of B-cell-specific gene expression), dead ringer (involved in development), and MRF-2 (which represses expression from the cytomegaloviris enhancer) (Dallas, P. B. et al. (2000) Mol. Cell Biol. 20:3137-3146). The ELM2 (Eg1-27 and MTA1 homology 2) domain is found in metastasis-associated protein MTA1 and protein ER1. The Caenorhabditis elegans gene eg1-27 is required for embryonic patterning MTA1, a human gene with elevated expression in metastatic carcinomas, is a component of a protein complex with histone deacetylase and nucleosome remodelling activities (Solari, F. et al. (1999) Development 126:2483-2494). The ELM2 domain is usually found to the N terminus of a myb-like DNA binding domain. ELM2 is also found associated with an ARID DNA. The Iroquois (Irx) family of genes are found in nematodes, insects and vertebrates. Irx genes usually occur in one or two genomic clusters of three genes each and encode transcriptional controllers that possess a characteristic homeodomain. The Irx genes function early in development to specify the identity of diverse territories of the body. Later in development in both Drosophila and vertebrates, the Irx genes function again to subdivide those territories into smaller domains. (For a review of Iroquois genes, see Cavodeassi, F. et al. (2001) Development 128:2847-2855.) For example, mouse and human Irx4 proteins are 83% conserved and their 63-aa homeodomain is more than 93% identical to that of the Drosophila Iroquois patterning genes. Irx4 transcripts are predominantly expressed in the cardiac ventricles. The homeobox gene Irx4 mediates ventricular differentiation during cardiac development (Bruneau, B. G. et al. (2000) Dev. Biol. 217:266-77). Histidine triad (HIT) proteins share residues in distinctive dimeric, 10-stranded half-barrel structures that form two identical purine nucleotide-binding sites. Hint (histidine triad nucleotide-binding protein)-related proteins, found in all forms of life, and fragile histidine triad (Fhit)-related proteins, found in animals and fungi, represent the two main branches of the HIT superfamily. Fhit homologs bind and cleave diadenosine polyphosphates. Fhit-Ap(n)A complexes appear to function in a proapoptotic tumor suppression pathway in epithelial tissues (Brenner C. et al. (1999) J. Cell Physiol.181:179-187). Most transcription factors contain characteristic DNA binding motifs, and variations on the above motifs and new motifs have been and are currently being characterized (Faisst, S. and S. Meyer (1992) Nucleic Acids Res. 20:3-26). These include the forkhead motif, found in transcription factors involved in development and oncogenesis (Hacker, U. et al. (1995) EMBO J 14:5306-5317), and the T-box protein T-domain, which forms a novel major and minor groove DNA contact T-box genes such as Brachyury (T) are essential for tissue specification in development (Muller, C. W. and B. G. Herrmann (1997) Nature 389:884-888). Mga is a novel protein which interacts with Max, a small bHLHZip protein required by Myc, Mad, and Mnt proteins to function as transcription factors. Max is required of these proteins for specific DNA binding to E-box sequences. Mga, like Myc, contains the basic-helix-loop-helix-leucine zipper motif (bHLHZip) and requires heterodimerization with Max for binding to the preferred Myc-Max-binding site CACGTG, but otherwise shows no relationship with Myc, Mad, or Mnt proteins. Mga also contains a DNA-binding domain called a T-box or T-domain. The T-domain, a highly conserved DNA-binding motif originally defined in the gastrulation-associated gene, Brachyury, is characteristic of the Tbx family of transcription factors. Mga binds the preferred Brachyury -binding sequence and represses transcription of reporter genes containing promoter-proximal Brachyury -binding sites. Mga is converted to a transcription activator of both Myc-Max and Brachyury site-containing reporters in a Max-dependent manner. Mga apparently functions as a dual-specificity transcription factor that regulates the expression of both Max-network and T-box family target genes (Hurlin, P. J. et al. (1999) EMBO J. 18:7019-7028). PGC-1 stands for thermogenic peroxisome proliferator-activated receptor gamma (PPAR-gamma) coactivator 1. It activates mitochondrial biogenesis in part through a direct interaction with nuclear respiratory factor 1 (NRF-1). A functional relative, PRC (PGC-1-related coactivator) is ubiquitously expressed in murine and human tissues and cell lines; but unlike PGC-1, PRC is not dramatically up-regulated during thermogenesis in brown fat. Its expression is down-regulated in quiescent BALB/3T3 cells and is rapidly induced by reintroduction of serum, conditions where PGC-1 is not detected. Similar to PGC-1, PRC activates NRF-1-dependent promoters. PRC interacts in vitro with the NRF-1 DNA binding domain through two distinct recognition motifs that are separated by an unstructured proline-rich region. PRC also contains a potent transcriptional activation domain in its amino terminus adjacent to an LXXLL motif. The spatial arrangement of these functional domains coincides with those found in PGC-1 (Andersson, U. and Scarpulla, R. C. (2001) Mol. Cell. Biol. 21:3738-3749). Chromatin Associated Proteins In the nucleus, DNA is packaged into chromatin, the compact organization of which limits the accessibility of DNA to transcription factors and plays a key role in gene regulation (Lewin, supra, pp. 409-410). The compact structure of chromatin is determined and influenced by chromatin-associated proteins such as the histones, the high mobility group (HMG) proteins, and the chromodomain proteins. There are five classes of histones, H1, H2A, H2B, H3, and H4, all of which are highly basic, low molecular weight proteins. The fundamental unit of chromatin, the nucleosome, consists of 200 base pairs of DNA associated with two copies each of H2A, H2B, H3, and H4. H1 links adjacent nucleosomes. HMG proteins are low molecular weight, non-histone proteins that may play a role in unwinding DNA and stabilizing single-stranded DNA. Chromodomain proteins play a key role in the formation of highly compacted heterochromatin, which is transcriptionally silent. Protamines are small, highly basic proteins that substitute for histones in sperm chromatin during the haploid phase of spermatogenesis. They pack sperm DNA into a highly condensed, stable, and inactive complex (Prosite PDOC00047 Protamine P1 signature). Higher-order structures of chromosomes involve the interaction of histones and chromosomal DNA with a series of nonhistone proteins. For example, HIRA is a histone binding protein that is a major candidate for causing developmental disorders associated with deletions in chromosome 22, including DiGeorge syndrome and velocardiofacial syndrome. HIRA interacts with core histones as well as the HIRA interacting protein HIRIP3 to form a complex that may have a role in regulating chromatin structure during development (Lorain, S. et al. (1998) Mol. Cell. Biol. 18:5546-5556). Diseases and Disorders Related to Gene Regulation Mutations in transcription factors contribute to oncogenesis. This is likely due to the role of transcription factors in the expression of genes involved in cell proliferation. For example, mutations in transcription factors encoded by proto-oncogenes, such as Fos, Jun, Myc, Rel, and Spi1, may be oncogenic due to increased stimulation of cell proliferation. Conversely, mutations in transcription factors encoded by tumor suppressor genes, such as p53, RB1, and WT1, may be oncogenic due to decreased inhibition of cell proliferation (Latchman, D. (1995) Gene Regulation: A Eukaryotic Perspective, Chapman and Hall, London, UK, pp. 242-255). Many neoplastic disorders in humans can be attributed to inappropriate gene expression. Malignant cell growth may result from either excessive expression of tumor promoting genes or insufficient expression of tumor suppressor genes (Cleary, M. L. (1992) Cancer Surv. 15:89-104). The zinc finger-type transcriptional regulator WT1 is a tumor-suppressor protein that is inactivated in children with Wilm's tumor. Deletions of the WT1 gene, or point mutations which destroy the DNA-binding activity of the protein, are associated with development of the pediatric nephroblastoma, Wilms tumor, and Denys-Drash syndrome (Rauscher, F. J. (1993) FASEB J. 7:896-903). The oncogene bcl-6, which plays an important role in large-cell lymphoma, is also a zinc-finger protein (Papavassiliou, A. G. (1995) N. Engl. J. Med. 332:45-47). Chromosomal translocations may also produce chimeric loci that fuse the coding sequence of one gene with the regulatory regions of a second unrelated gene. Such an arrangement likely results in inappropriate gene transcription, potentially contributing to malignancy. In Burkitt's lymphoma, for example, the transcription factor Myc is translocated to the immunoglobulin heavy chain locus, greatly enhancing Myc expression and resulting in rapid cell growth leading to leukemia (Latchman, D. S. (1996) N. Engl. J. Med. 334:28-33). Human acute leukemias involve reciprocal chromosome translocations that fuse the ALL-1 gene located at chromosome region 11q23 to a series of partner genes positioned on a variety of human chromosomes. The fused genes encode chimeric proteins. The AF17 gene encodes a protein of 1093 amino acids, containing a leucine-zipper dimerization motif located 3′ of the fusion point and a cysteine-rich domain at the N terminus that shows homology to a domain within the protein Br140 (peregrin) (Prasad R. et al. (1994) Proc. Natl. Acad. Sci. USA 91:8107-8111). Certain proteins enriched in glutamine are associated with various neurological disorders including spinocerebellar ataxia, bipolar effective disorder, schizophrenia, and autism (Margolis, R. L. et al. (1997) Human Genetics 100:114-122). These proteins contain regions with as many as 15 or more consecutive glutamine residues and may function as transcription factors with a potential role in regulation of neurodevelopment or neuroplasticity. Impaired transcriptional regulation may lead to Alzheimer's disease, a progressive neurodegenerative disorder that is characterized by the formation of senile plaques and neurofibrillary tangles containing amyloid beta peptide. These plaques are found in limbic and association cortices of the brain, including hippocampus, temporal cortices, cingulate cortex, amygdala, nucleus basalis and locus caeruleus. Early in Alzheimer's pathology, physiological changes are visible in the cingulate cortex (Minoshima, S. et al. (1997) Ann. Neurol. 42:85-94). In subjects with advanced Alzheimer's disease, accumulating plaques damage the neuronal architecture in limbic areas and eventually cripple the memory process. In addition, the immune system responds to infection or trauma by activating a cascade of events that coordinate the progressive selection, amplification, and mobilization of cellular defense mechanisms. A complex and balanced program of gene activation and repression is involved in this process. However, hyperactivity of the immune system as a result of improper or insufficient regulation of gene expression may result in considerable tissue or organ damage. This damage is well-documented in immunological responses associated with arthritis, allergens, heart attack, stroke, and infections. (Isselbacher, K. J. et al. Harrison's Principles of Internal Medicine, 13/e, McGraw Hill, Inc. and Teton Data Systems Software, 1996.) In particular, a zinc finger protein termed Staf50 (for Stimulated trans-acting factor of 50 kDa) is a transcriptional regulator and is induced in various cell lines by interferon-I and -II. Staf50 appears to mediate the antiviral activity of interferon by down-regulating the viral transcription directed by the long terminal repeat promoter region of human immunodeficiency virus type-1 in transfected cells (Tissot, C. (1995) J. Biol. Chem. 270:14891-14898). Also, the causative gene for autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) was recently isolated and found to encode a protein with two PHD-type zinc finger motifs (Bjorses, P. et al. (1998) Hum. Mol. Genet. 7:1547-1553). Furthermore, the generation of multicellular organisms is based upon the induction and coordination of cell differentiation at the appropriate stages of development. Central to this process is differential gene expression, which confers the distinct identities of cells and tissues throughout the body. Failure to regulate gene expression during development could result in developmental disorders. Human developmental disorders caused by mutations in zinc finger-type transcriptional regulators include: urogenital developmental abnormalities associated with WT1; Greig cephalopolysyndactyly, Pallister-Hall syndrome, and postaxial polydactyly type A (GLI3), and Townes-Brocks syndrome, characterized by anal, renal, limb, and ear abnormalities (SALL1) (Engelkamp, D. and V. van Heyningen (1996) Curr. Opin. Genet. Dev. 6:334-342; Kohlhase, J. et al. (1999) Am. J. Hum. Genet. 64:435-445). Synthesis of Nucleic Acids Polymerases DNA and RNA replication are critical processes for cell replication and function. DNA and RNA replication are mediated by the enzymes DNA and RNA polymerase, respectively, by a “templating” process in which the nucleotide sequence of a DNA or RNA strand is copied by complementary base-pairing into a complementary nucleic acid sequence of either DNA or RNA. However, there are fundamental differences between the two processes. DNA polymerase catalyzes the stepwise addition of a deoxyribonucleotide to the 3′-OH end of a polynucleotide strand (the primer strand) that is paired to a second (template) strand. The new DNA strand therefore grows in the 5′ to 3′ direction (Alberts, et al.,supra, pp. 251-254). The substrates for the polymerization reaction are the corresponding deoxynucleotide triphosphates which must base-pair with the correct nucleotide on the template strand in order to be recognized by the polymerase. Because DNA exists as a double-stranded helix, each of the two strands may serve as a template for the formation of a new complementary strand. Each of the two daughter cells of a dividing cell therefore inherits a new DNA double helix containing one old and one new strand. Thus, DNA is said to be replicated “semiconservatively” by DNA polymerase. In addition to the synthesis of new DNA, DNA polymerase is also involved in the repair of damaged DNA as discussed below under “Ligases.” In contrast to DNA polymerase, RNA polymerase uses a DNA template strand to “transcribe” DNA into RNA using ribonucleotide triphosphates as substrates. Like DNA polymerization, RNA polymerization proceeds in a 5′ to 3′ direction by addition of a ribonucleoside monophosphate to the 3′-OH end of a growing RNA chain. DNA transcription generates messenger RNAs (mRNA) that carry information for protein synthesis, as well as the transfer, ribosomal, and other RNAs that have structural or catalytic functions. In eukaryotes, three discrete RNA polymerases synthesize the three different types of RNA (Alberts et al., supra, pp. 367-368). RNA polymerase I makes the large ribosomal RNAs, RNA polymerase II makes the mRNAs that will be translated into proteins, and RNA polymerase III makes a variety of small, stable RNAs, including 5S ribosomal RNA and the transfer RNAs (tRNA). In all cases, RNA synthesis is initiated by binding of the RNA polymerase to a promoter region on the DNA and synthesis begins at a start site within the promoter. Synthesis is completed at a stop (termination) signal in the DNA whereupon both the polymerase and the completed RNA chain are released. Ligases DNA repair is the process by which accidental base changes, such as those produced by oxidative damage, hydrolytic attack, or uncontrolled methylation of DNA, are corrected before replication or transcription of the DNA can occur. Because of the efficiency of the DNA repair process, fewer than one in a thousand accidental base changes causes a mutation (Alberts et al., supra, pp. 245-249). The three steps common to most types of DNA repair are (1) excision of the damaged or altered base or nucleotide by DNA nucleases, (2) insertion of the correct nucleotide in the gap left by the excised nucleotide by DNA polymerase using the complementary strand as the template and, (3) sealing the break left between the inserted nucleotide(s) and the existing DNA strand by DNA ligase. In the last reaction, DNA ligase uses the energy from ATP hydrolysis to activate the 5′ end of the broken phosphodiester bond before forming the new bond with the 3′-OH of the DNA strand. In Bloom's syndrome, an inherited human disease, individuals are partially deficient in DNA ligation and consequently have an increased incidence of cancer (Alberts et al., supra, p. 247). Nucleases Nucleases comprise enzymes that hydrolyze both DNA (DNase) and RNA (Rnase). They serve different purposes in nucleic acid metabolism. Nucleases hydrolyze the phosphodiester bonds between adjacent nucleotides either at internal positions (endonucleases) or at the terminal 3′ or 5′ nucleotide positions (exonucleases). A DNA exonuclease activity in DNA polymerase, for example, serves to remove improperly paired nucleotides attached to the 3′-OH end of the growing DNA strand by the polymerase and thereby serves a “proofreading” function. As mentioned above, DNA endonuclease activity is involved in the excision step of the DNA repair process. RNases also serve a variety of functions. For example, RNase P is a ribonucleoprotein enzyme which cleaves the 5′ end of pre-tRNAs as part of their maturation process. RNase H digests the RNA strand of an RNA/DNA hybrid. Such hybrids occur in cells invaded by retroviruses, and RNase H is an important enzyme in the retroviral replication cycle. Pancreatic RNase secreted by the pancreas into the intestine hydrolyzes RNA present in ingested foods. RNase activity in serum and cell extracts is elevated in a variety of cancers and infectious diseases (Schein, C. H. (1997) Nat. Biotechnol. 15:529-536). Regulation of RNase activity is being investigated as a means to control tumor angiogenesis, allergic reactions, viral infection and replication, and fungal infections. Modification of Nucleic Acids DNA Repair Cells are constantly faced with replication errors and environmental assault (such as ultraviolet irradiation) that can produce DNA damage. Damage to DNA consists of any change that modifies the structure of the molecule. Changes to DNA can be divided into two general classes, single base changes and structural distortions. Single base changes affect the sequence but not the overall structure of the DNA. Since single base changes do not affect transcription or replication, they exert their effect on future generations. Structural distortions affect the structure of the DNA. A single strand nick or removal of a base may prevent a strand from acting as a viable template for synthesis of DNA or RNA. Intrastrand or interstrand covalent linkage between bases, or the addition of a bulky adduct to a base, may distort the structure of the double helix and interfere with transcription and replication. Any damage to DNA can produce a mutation, and the mutation may produce a disorder, such as cancer. Changes in DNA are recognized by repair systems within the cell. These repair systems act to correct the damage and thus prevent any deleterious affects of a mutational event. Repair systems can be divided into three general types, direct repair, excision repair, and retrieval systems. When the repair systems are eliminated, cells become exceedingly sensitive to environmental mutagens, such as ultraviolet irradiation. Disorders associated with a loss in DNA repair systems often exhibit a high sensitivity to environmental mutagens. Examples of such disorders include xeroderma pigmentosum, Bloom's syndrome, and Werner's syndrome. Xeroderma pigmentosum results in a hypersensitivity to sunlight, especially ultraviolet, and produces skin defects. Bloom's syndrome results in an increased frequency of chromosomal aberrations, including sister chromosome exchanges (Yamagata, K. et al. (1998) Proc. Natl. Acad. Sci. USA 95:8733-8738). Direct repair involves the reversal or simple removal of the damaged region of DNA. Mismatches involving normal bases are repaired based on certain biases within the repair system. For example, mismatched GT base pairs are frequently caused by deamination of 5-methyl-cytosine to form thymine. Therefore, repair systems convert mismatched GT pairs to GC, instead of AT. Repair also favors the non-methylated strand in hemimethylated DNA, since this strand represents the newly synthesized daughter strand. The recognition of hemimethylated DNA and repair of mismatches on the non-methylated strand involve the products of the genes mutH, mutL, mutS (which specifically recognizes mismatched base pairs), the helicase encoded by the uvrD gene, and the methylase encoded by the dam gene. C-5 cytosine-specific DNA methylases are enzymes that specifically methylate the C-5 carbon of cytosines in DNA (Kumar, S. et al. (1994) Nucleic Acids Res. 22:1-10). Excision repair is a system in which mispaired or damaged bases are removed from DNA and a new stretch of DNA is synthesized to replace them In the incision step, the damaged structure is recognized by an endonuclease that cleaves the DNA strand on both sides of the damage. In the excision step, a 5′-3′exonuclease removes a stretch of the damaged DNA strand. In the synthesis step, the resulting single-stranded region serves as a template for a DNA polymerase to synthesize a replacement for the excised sequence. Finally, DNA ligase covalently links the 3′ end of the new material to the old material. In mammals, DNA polymerase beta serves as the DNA repair polymerase. Mutations in the human DNA polymerase beta gene are associated with several types of cancer (Bhattacharyya, N. et al. (1999) DNA Cell Biol. 18:549-554; Matsuzaki, J. et al. (1996) Mol. Carcinog. 15:38-43). Methylases Methylation of specific nucleotides occurs in both DNA and RNA, and serves different functions in the two macromolecules. Methylation of cytosine residues to form 5-methyl cytosine in DNA occurs specifically in CG sequences which are base-paired with one another in the DNA double-helix. The pattern of methylation is passed from generation to generation during DNA replication by an enzyme called “maintenance methylase” that acts preferentially on those CG sequences that are base-paired with a CG sequence that is already methylated. Such methylation appears to distinguish active from inactive genes by preventing the binding of regulatory proteins that “turn on” the gene, but permiting the binding of proteins that inactivate the gene (Alberts et al., supra, pp. 448-451). N-6 adenine-specific methylases are enzymes that specifically methylate the amino group at the C-6 position of adenines in DNA. These enzymes are found in the three known types of bacterial restriction-modification, systems (Prosite PDOC00087 N-6 Adenine-specific DNA methylases signature). In RNA metabolism, “tRNA methylase” produces one of several nucleotide modifications in tRNA that affect the conformation and base-pairing of the molecule and facilitate the recognition of the appropriate mRNA codons by specific tRNAs. The primary methylation pattern is the dimethylation of guanine residues to form N,N-dimethyl guanine. Helicases and Single-stranded Binding Proteins Helicases are enzymes that destabilize and unwind double helix structures in both DNA and RNA. Since DNA replication occurs more or less simultaneously on both strands, the two strands must first separate to generate a replication “fork” for DNA polymerase to act on. Two types of replication proteins contribute to this process, DNA helicases and single-stranded binding proteins. DNA helicases hydrolyze ATP and use the energy of hydrolysis to separate the DNA strands. Single-stranded binding proteins (SSBs) then bind to the exposed DNA strands, without covering the bases, thereby temporarily stabilizing them for templating by the DNA polymerase (Alberts et al., supra, pp. 255-256). RNA helicases also alter and regulate RNA conformation and secondary structure. Like the DNA helicases, RNA helicases utilize energy derived from ATP hydrolysis to destabilize and unwind RNA duplexes. The most well-characterized and ubiquitous family of RNA helicases is the DEAD-box family, so named for the conserved B-type ATP-binding motif which is diagnostic of proteins in this family. Over 40 DEAD-box helicases have been identified in organisms as diverse as bacteria, insects, yeast, amphibians, mammals, and plants. DEAD-box helicases function in diverse processes such as translation initiation, splicing, ribosome assembly, and RNA editing, transport, and stability. Examples of these RNA helicases include yeast Drs1 protein, which is involved in ribosomal RNA processing; yeast TIF1 and TIF2 and mammalian eIF-4A, which are essential to the initiation of RNA translation; and human p68 antigen, which regulates cell growth and division (Ripmaster, T. L. et al. (1992) Proc. Natl. Acad. Sci. USA 89:11131-11135; Chang, T.-H. et al. (1990) Proc. Natl. Acad. Sci. USA 87:1571-1575). These RNA helicases demonstrate strong sequence homology over a stretch of some 420 amino acids. Included among these conserved sequences are the consensus sequence for the A motif of an ATP binding protein; the “DEAD box” sequence, associated with ATPase activity; the sequence SAT, associated with the actual helicase unwinding region; and an octapeptide consensus sequence, required for RNA binding and ATP hydrolysis (Pause, A. et al. (1993) Mol. Cell Biol. 13:6789-6798). Differences outside of these conserved regions are believed to reflect differences in the functional roles of individual proteins (Chang et al., supra). Some DEAD-box helicases play tissue- and stage-specific roles in spermatogenesis and embryogenesis. Overexpression of the DEAD-box 1 protein (DDX1) may play a role in the progression of neuroblastoma (Nb) and retinoblastoma (Rb) tumors (Godbout, R. et al. (1998) J. Biol. Chem. 273:21161-21168). These observations suggest that DDX1 may promote or enhance tumor progression by altering the normal secondary structure and expression levels of RNA in cancer cells. Other DEAD-box helicases have been implicated either directly or indirectly in tumorigenesis (Godbout et al., supra). For example, murine p68 is mutated in ultraviolet light-induced tumors, and human DDX6 is located at a chromosomal breakpoint associated with B-cell lymphoma. Similarly, a chimeric protein comprised of DDX10 and NUP98, a nucleoporin protein, may be involved in the pathogenesis of certain myeloid malignancies. Topoisomerases Besides the need to separate DNA strands prior to replication, the two strands must be “unwound” from one another prior to their separation by DNA helicases. This function is performed by proteins known as DNA topoisomerases. DNA topoisomerase effectively acts as a reversible nuclease that hydrolyzes a phosphodiesterase bond in a DNA strand, permits the two strands to rotate freely about one another to remove the strain of the helix, and then rejoins the original phosphodiester bond between the two strands. Topoisomerases are essential enzymes responsible for the topological rearrangement of DNA brought about by transcription, replication, chromatin formation, recombination, and chromosome segregation. Superhelical coils are introduced into DNA by the passage of processive enzymes such as RNA polymerase, or by the separation of DNA strands by a helicase prior to replication. Knotting and concatenation can occur in the process of DNA synthesis, storage, and repair. All topoisomerases work by breaking a phosphodiester bond in the ribose-phosphate backbone of DNA. A catalytic tyrosine residue on the enzyme makes a nucleophilic attack on the scissile phosphodiester bond, resulting in a reaction intermediate in which a covalent bond is formed between the enzyme and one end of the broken strand. A tyrosine-DNA phosphodiesterase functions in DNA repair by hydrolyzing this bond in occasional dead-end topoisomerase I-DNA intermediates (Pouliot, J. J. et al. (1999) Science 286:552-555). Two types of DNA topoisomerase exist, types I and II. Type I topoisomerases work as monomers, making a break in a single strand of DNA while type II topoisomerases, working as homodimers, cleave both strands. DNA Topoisomerase I causes a single-strand break in a DNA helix to allow the rotation of the two strands of the helix about the remaining phosphodiester bond in the opposite strand. DNA topoisomerase II causes a transient break in both strands of a DNA helix where two double helices cross over one another. This type of topoisomerase can efficiently separate two interlocked DNA circles (Alberts et al., supra, pp. 260-262). Type II topoisomerases are largely confined to proliferating cells in eukaryotes, such as cancer cells. For this reason they are targets for anticancer drugs. Topoisomerase II has been implicated in multi-drug resistance (MDR) as it appears to aid in the repair of DNA damage inflicted by DNA binding agents such as doxorubicin and vincristine. The type II topoisomerases are specific targets of drug classes that comprise complex-stabilizing (epipodophyllotoxins, anthracyclines) and catalytic (merbarone, bisdioxopiperazines) inhibitors (Beck, W. T. et al. (1999) Drug Resist. Update 2:382-389). Topoisomerases include topo IIalpha-1 and topo IIbeta-1; topo IIalpha-2 and topo IIbeta-2, are novel variants that appear to be conserved between chicken and human. Topo IIalpha-2 encodes a protein with an additional 35 amino acids inserted after K321 of the chicken topo IIalpha-1 protein sequence. Topo IIbeta-2 encodes a protein missing 86 amino acids following V27 in the topo IIbeta-1 protein sequence. Alternatively spliced forms of human topo IIalpha are also observed (Petruti-Mot, A. S. and Earnshaw, W. C. (2000) Gene 258:183-192). The topoisomerase I family includes topoisomerases I and III (topo I and topo III). The crystal structure of human topoisomerase I suggests that rotation about the intact DNA strand is partially controlled by the enzyme. In this “controlled rotation” model, protein-DNA interactions limit the rotation, which is driven by torsional strain in the DNA (Stewart, L. et al. (1998) Science 379:1534-1541). Structurally, topo I can be recognized by its catalytic tyrosine residue and a number of other conserved residues in the active site region. Topo I is thought to function during transcription. Two topo IIIs are known in humans, and they are homologous to prokaryotic topoisomerase I, with a conserved tyrosine and active site signature specific to this family. Topo III has been suggested to play a role in meiotic recombination. A mouse topo III is highly expressed in testis tissue and its expression increases with the increase in the number of cells in pachytene (Seki, T. et al. (1998) J. Biol. Chem. 273:28553-28556). The topoisomerase II family includes two isozymes (IIα and IIβ) encoded by different genes. Topo II cleaves double stranded DNA in a reproducible, nonrandom fashion, preferentially in an AT rich region, but the basis of cleavage site selectivity is not known. Structurally, topo II is made up of four domains, the first two of which are structurally similar and probably distantly homologous to similar domains in eukaryotic topo I. The second domain bears the catalytic tyrosine, as well as a highly conserved pentapeptide. The IIα isoform appears to be responsible for unlinking DNA during chromosome segregation. Cell lines expressing IIα but not IIβ suggest that IIβ is dispensable in cellular processes; however, IIβ knockout mice died perinatally due to a failure in neural development. That the major abnormalities occurred in predominantly late developmental events (neurogenesis) suggests that IIβ is needed not at mitosis, but rather during DNA repair (Yang, X. et al. (2000) Science 287:131-134). Topoisomerases have been implicated in a number of disease states, and topoisomerase poisons have proven to be effective anti-tumor drugs for some human malignancies. Topo I is mislocalized in Fanconi's anemia, and may be involved in the chromosomal breakage seen in this disorder (Wunder, E. (1984) Hum. Genet. 68:276-281). Overexpression of a truncated topo III in ataxia-telangiectasia (A-T) cells partially suppresses the A-T phenotype, probably through a dominant negative mechanism. This suggests that topo III is deregulated in A-T (Fritz, E. et al. (1997) Proc. Natl. Acad. Sci. USA 94:4538-4542). Topo III also interacts with the Bloom's Syndrome gene product, and has been suggested to have a role as a tumor suppressor (Wu, L. et al. (2000) J. Biol. Chem 275:9636-9644). Aberrant topo II activity is often associated with cancer or increased cancer risk Greatly lowered topo II activity has been found in some, but not all A-T cell lines (Mohamed, R. et al. (1987) Biochem. Biophys. Res. Commun. 149:233-238). On the other hand, topo II can break DNA in the region of the A-T gene (ATM), which controls all DNA damage-responsive cell cycle checkpoints (Kaufmann, W. K. (1998) Proc. Soc. Exp. Biol. Med. 217:327-334). The ability of topoisomerases to break DNA has been used as the basis of antitumor drugs. Topoisomerase poisons act by increasing the number of dead-end covalent DNA-enzyme complexes in the cell, ultimately triggering cell death pathways (Fortune, J. M. and N. Osheroff (2000) Prog. Nucleic Acid Res. Mol. Biol. 64:221-253; Guichard, S. M. and M. K. Danks (1999) Curr. Opin. Oncol. 11:482-489). Antibodies against topo I are found in the serum of systemic sclerosis patients, and the levels of the antibody may be used as a marker of pulmonary involvement in the disease (Diot, E. et al. (1999) Chest 116:715-720). Finally, the DNA binding region of human topo I has been used as a DNA delivery vehicle for gene therapy (Chen, T. Y. et al. (2000) Appl. Microbiol. Biotechnol. 53:558-567). Recombinases Genetic recombination is the process of rearranging DNA sequences within an organism's genome to provide genetic variation for the organism in response to changes in the environment. DNA recombination allows variation in the particular combination of genes present in an individual's genome, as well as the timing and level of expression of these genes (Alberts et al., supra, pp. 263-273). Two broad classes of genetic recombination are commonly recognized, general recombination and site-specific recombination. General recombination involves genetic exchange between any homologous pair of DNA sequences usually located on two copies of the same chromosome. The process is aided by enzymes, recombinases, that “nick” one strand of a DNA duplex more or less randomly and permit exchange with a complementary strand on another duplex. The process does not normally change the arrangement of genes in a chromosome. In site-specific recombination, the recombinase recognizes specific nucleotide sequences present in one or both of the recombining molecules. Base-pairing is not involved in this form of recombination and therefore it does not require DNA homology between the recombining molecules. Unlike general recombination, this form of recombination can alter the relative positions of nucleotide sequences in chromosomes. RNA Metabolism Much of the regulation of gene expression in eucaryotic cells occurs at the posttranscriptional level. Messenger RNAs (mRNA), which are produced in the cell nucleus from primary transcripts of protein-encoding genes, are processed and transported to the cytoplasm where the protein synthesis machinery is located. RNA-binding proteins are a group of proteins that participate in the processing, editing, transport, localization, and posttranscriptional regulation of mRNAs, and comprise the protein component of ribosomes as well. The RNA-binding activity of many of these proteins is mediated by a series of RNA-binding motifs identified within them. These domains include the RNP motif, the arginine-rich motif, the RGG box, and the KH motif (Burd, C. G. and G. Dreyfuss (1994) Science 265:615-621). The RNP motif is the most widely found and best characterized of these motifs. The RNP motif is composed of 90-100 amino acids which form an RNA-binding domain and is found in one or more copies in proteins that bind pre-mRNA, mRNA, pre-ribosomal RNA, and small nuclear RNAs. The RNP motif is composed of two short sequences (RNP-1 and RNP-2) and a number of other mostly hydrophobic, conserved amino acids interspersed throughout the motif (Burd and Dreyfuss, supra; ExPASy PROSITE document PDOC0030). Ribonucleic acid (RNA) is a linear single-stranded polymer of four nucleotides, ATP, CTP, UTP, and GTP. In most organisms, RNA is transcribed as a copy of deoxyribonucleic acid (DNA), the genetic material of the organism. In retroviruses RNA rather than DNA serves as the genetic material. RNA copies of the genetic material encode proteins or serve various structural, catalytic, or regulatory roles in organisms. RNA is classified according to its cellular localization and function. Messenger RNAs (mRNAs) encode polypeptides. Ribosomal RNAs (rRNAs) are assembled, along with ribosomal proteins, into ribosomes, which are cytoplasmic particles that translate mRNA into polypeptides. Transfer RNAs (tRNAs) are cytosolic adaptor molecules that function in mRNA translation by recognizing both an mRNA codon and the amino acid that matches that codon. Heterogeneous nuclear RNAs (hnRNAs) include mRNA precursors and other nuclear RNAs of various sizes. Small nuclear RNAs (snRNAs) are a part of the nuclear spliceosome complex that removes intervening, non-coding sequences (introns) and rejoins exons in pre-mRNAs. Proteins are associated with RNA during its transcription from DNA, RNA processing, and translation of mRNA into protein. Proteins are also associated with RNA as it is used for structural, catalytic, and regulatory purposes. Transcription Transcription in eukaryotes is catalyzed by three species of RNA polymerase: RNA polymerase I for rRNA synthesis, RNA polymerase II for mRNA synthesis and RNA polymerase III for tRNA and 5S rRNA synthesis. Each RNA polymerase is composed of more than 10 different polypeptides. The RNA polymerase III enzymes are the most complex of the nuclear polymerases. They contain the largest number of subunits; their basal transcription machinery includes the core transcription factors (TF) IIIA, IIIB and IIIC; and they have promoters that are mostly located within transcribed DNA (Akira Ishihama et al. (1998) Curr. Opin. Microbiol. 1:190-196). cDNA and genomic clones have been isolated for the second-largest subunit of RNA polymerase III in Drosophila melanogaster. The deduced polypeptide, named DmRP128, consists of 1135 amino acids with a calculated molecular weight of 128 kDa. The protein sequence shares conserved regions of homology with other cloned the second-largest subunits of RNA polymerases (Seifarth, W. et al. (1991) Mol. Gen. Genet. 228:424-432). RNA Processing Ribosomal RNAs (rRNAs) are assembled, along with ribosomal proteins, into ribosomes, which are cytoplasmic particles that translate messenger RNA (mRNA) into polypeptides. The eukaryotic ribosome is composed of a 60S (large) subunit and a 40S (small) subunit, which together form the 80S ribosome. In addition to the 18S, 28S, 5S, and 5.8S rRNAs, ribosomes contain from 50 to over 80 different ribosomal proteins, depending on the organism. Ribosomal proteins are classified according to which subunit they belong (i.e., L, if associated with the large 60S large subunit or S if associated with the small 40S subunit). E. coli ribosomes have been the most thoroughly studied and contain 50 proteins, many of which are conserved in all life forms. The structures of nine ribosomal proteins have been solved to less than 3.0D resolution (i.e., S5, S6, S17, L1, L6, L9, L12, L14, L30), revealing common motifs, such as b-a-b protein folds in addition to acidic and basic RNA-binding motifs positioned between b-strands. Most ribosomal proteins are believed to contact rRNA directly (reviewed in Liljas, A. and M. Garber (1995) Curr. Opin. Struct. Biol. 5:721-727; see also Woodson, S. A. and N. B. Leontis (1998) Curr. Opin. Struct. Biol. 8:294-300; Ramakrishnan, V. and S. W. White (1998) Trends Biochem. Sci. 23:208-212). Ribosomal proteins may undergo post-translational modifications or interact with other ribosome-associated proteins to regulate translation. For example, the highly homologous 40S ribosomal protein S6 kinases (S6K1 and S6K2) play a key role in the regulation of cell growth by controlling the biosynthesis of translational components which make up the protein synthetic apparatus (including the ribosomal proteins). In the case of S6K1, at least eight phosphorylation sites are believed to mediate kinase activation in a hierarchical fashion (Dufner, A and G. Thomas (1999) Exp. Cell. Res. 253:100-109). Some of the ribosomal proteins, including L1, also function as translational repressors by binding to polycistronic mRNAs encoding ribosomal proteins (Liljas, supra and Garber, supra). Recent evidence suggests that a number of ribosomal proteins have secondary functions independent of their involvement in protein biosynthesis. These proteins functions as regulators of cell proliferation and, in some instances, as inducers of cell death. For example, the expression of human ribosomal protein L13a has been shown to induce apoptosis by arresting cell growth in the G2/M phase of the cell cycle. Inhibition of expression of L13a induces apoptosis in target cells, which suggests that this protein is necessary, in the appropriate amount, for cell survival. Similar results have been obtained in yeast where inactivation of yeast homologues of L13a, rp22 and rp23, results in severe growth retardation and death. A closely related ribosomal protein, L7, arrests cells in G1 and also induces apoptosis. Thus, it appears that a subset of ribosomal proteins may function as cell cycle checkpoints and compose a new family of cell proliferation regulators. Mapping of individual ribosomal proteins on the surface of intact ribosomes is accomplished using 3D immunocryoelectronmicroscopy, whereby antibodies raised against specific ribosomal proteins are visualized. Progress has been made toward the mapping of L1, L7, and L12 while the structure of the intact ribosome has been solved to only 20-25D resolution and inconsistencies exist among different crude structures (Frank, J. (1997) Curr. Opin. Struct. Biol. 7:266-272). Three distinct sites have been identified on the ribosome. The aminoacyl-tRNA acceptor site (A site) receives charged tRNAs (with the exception of the initiator-tRNA). The peptidyl-tRNA site (P site) binds the nascent polypeptide as the amino acid from the A site is added to the elongating chain. Deacylated tRNAs bind in the exit site (E site) prior to their release from the ribosome. (The structure of the ribosome is reviewed in Stryer, L. (1995) Biochemistry, W. H. Freeman and Company, New York N.Y., pp. 888-908; Lodish, supra, pp. 119-138; and Lewin, B. (1997) Genes VI, Oxford University Press, Inc. New York N.Y.). Various proteins are necessary for processing of transcribed RNAs in the nucleus. Pre-mRNA processing steps include capping at the 5′ end with methylguanosine, polyadenylating the 3′ end, and splicing to remove introns. The primary RNA transript from DNA is a faithful copy of the gene containing both exon and intron sequences, and the latter sequences must be cut out of the RNA transcript to produce a mRNA that codes for a protein. This “splicing” of the mRNA sequence takes place in the nucleus with the aid of a large, multicomponent ribonucleoprotein complex known as a spliceosome. The spliceosomal complex is comprised of five small nuclear ribonucleoprotein particles (snRNPs) designated U1, U2, U4, U5, and U6. Each snRNP contains a single species of snRNA and about ten proteins. The RNA components of some snRNPs recognize and base-pair with intron consensus sequences. The protein components mediate spliceosome assembly and the splicing reaction. Autoantibodies to snRNP proteins are found in the blood of patients with systemic lupus erythematosus (Stryer, supra, p. 863). Heterogeneous nuclear ribonucleoproteins (hnRNPs) have been identified that have roles in splicing, exporting of the mature RNAs to the cytoplasm, and mRNA translation (Biamonti, G. et al. (1998) Clin. Exp. Rheumatol. 16:317-326). Some examples of hnRNPs include the yeast proteins Hrp1p, involved in cleavage and polyadenylation at the 3′ end of the RNA; Cbp80p, involved in capping the 5′ end of the RNA; and Np13p, a homolog of mammalian hnRNP A1, involved in export of mRNA from the nucleus (Shen, E. C. et al. (1998) Genes Dev. 12:679-691). HnRNPs have been shown to be important targets of the autoimmune response in rheumatic diseases (Biamonti et al., supra). Many snRNP and hnRNP proteins are characterized by an RNA recognition motif (RRM) (reviewed in Birney, E. et al. (1993) Nucleic Acids Res. 21:5803-5816). The RRM is about 80 amino acids in length and forms four β-strands and two α-helices arranged in an α/β sandwich. The RRM contains a core RNP-1 octapeptide motif along with surrounding conserved sequences. In addition to snRNP proteins, examples of RNA-binding proteins which contain the above motifs include heteronuclear ribonucleoproteins which stabilize nascent RNA and factors which regulate alternative splicing. Alternative splicing factors include developmentally regulated proteins, specific examples of which have been identified in lower eukaryotes such as Drosophila melanogaster and Caenorhabditis elegans. These proteins play key roles in developmental processes such as pattern formation and sex determination, respectively (Hodgkin, J. et al. (1994) Development 120:3681-3689). The 3′ ends of most eukaryote mRNAs are also posttranscriptionally modified by polyadenylation. Polyadenylation proceeds through two enzymatically distinct steps: (i) the endonucleolytic cleavage of nascent mRNAs at cis-acting polyadenylation signals in the 3′-untranslated (non-coding) region and (ii) the addition of a poly(A) tract to the 5′ mRNA fragment. The presence of cis-acting RNA sequences is necessary for both steps. These sequences include 5′-AAUAAA-3′ located 10-30 nucleotides upstream of the cleavage site and a less well-conserved GU- or U-rich sequence element located 10-30 nucleotides downstream of the cleavage site. Cleavage stimulation factor (CstF), cleavage factor I (CF I), and cleavage factor II (CF II) are involved in the cleavage reaction while cleavage and polyadenylation specificity factor (CPSF) and poly(A) polymerase (PAP) are necessary for both cleavage and polyadenylation. An additional enzyme, poly(A)-binding protein II (PAB II), promotes poly(A) tract elongation (Rüegsegger, U. et al. (1996) J. Biol. Chem. 271:6107-6113; and references within). Translation Correct translation of the genetic code depends upon each amino acid forming a linkage with the appropriate transfer RNA (tRNA). The aminoacyl-tRNA synthetases (aaRSs) are essential proteins found in all living organisms. The aaRSs are responsible for the activation and correct attachment of an amino acid with its cognate tRNA, as the first step in protein biosynthesis. Prokaryotic organisms have at least twenty different types of aaRSs, one for each different amino acid, while eukaryotes usually have two aaRSs, a cytosolic form and a mitochondrial form, for each different amino acid. The 20 aaRS enzymes can be divided into two structural classes. Class I enzymes add amino acids to the 2′ hydroxyl at the 3′ end of tRNAs while Class II enzymes add amino acids to the 3′ hydroxyl at the 3′ end of tRNAs. Each class is characterized by a distinctive topology of the catalytic domain. Class I enzymes contain a catalytic domain based on the nucleotide-binding ‘Rossman fold’. In particular, a consensus tetrapeptide motif is highly conserved (Prosite Document PDOC00161, Aminoacyl-transfer RNA synthetases class-I signature). Class I enzymes are specific for arginine, cysteine, glutamic acid, glutamine, isoleucine, leucine, methionine, tyrosine, tryptophan, and valine. Class II enzymes contain a central catalytic domain, which consists of a seven-stranded antiparallel β-sheet domain, as well as N— and C-terminal regulatory domains. Class II enzymes are separated into two groups based on the heterodimeric or homodimeric structure of the enzyme; the latter group is further subdivided by the structure of the N— and C-terminal regulatory domains (Hartlein, M. and S. Cusack (1995) J. Mol. Evol. 40:519-530). Class II enzymes are specific for alanine, asparagine, aspartic acid, glycine, histidine, lysine, phenylalanine, proline, serine, and threonine. Certain aaRSs also have editing functions. IleRS, for example, can misactivate valine to form Val-tRNA Ile , but this product is cleared by a hydrolytic activity that destroys the mischarged product. This editing activity is located within a second catalytic site found in the connective polypeptide 1 region (CP1), a long insertion sequence within the Rossman fold domain of Class I enzymes (Schimmel, P. et al. (1998) FASEB J. 12:1599-1609). AaRSs also play a role in tRNA processing. It has been shown that mature tRNAs are charged with their respective amino acids in the nucleus before export to the cytoplasm, and charging may serve as a quality control mechanism to insure the tRNAs are functional (Martinis, S. A. et al. (1999) EMBO J. 18:4591-4596). Under optimal conditions, polypeptide synthesis proceeds at a rate of approximately 40 amino acid residues per second. The rate of misincorporation during translation in on the order of 10 −4 and is primarily the result of aminoacyl-t-RNAs being charged with the incorrect amino acid. Incorrectly charged tRNA are toxic to cells as they result in the incorporation of incorrect amino acid residues into an elongating polypeptide. The rate of translation is presumed to be a compromise between the optimal rate of elongation and the need for translational fidelity. Mathematical calculations predict that 10 −4 is indeed the maximum acceptable error rate for protein synthesis in a biological system (reviewed in Stryer, supra; and Watson, J. et al. (1987) The Benjamin/Cummings Publishing Co., Inc. Menlo Park, Calif.). A particularly error prone aminoacyl-tRNA charging event is the charging of tRNA Gln with Gln. A mechanism exits for the correction of this mischarging event which likely has its origins in evolution. Gln was among the last of the 20 naturally occurring amino acids used in polypeptide synthesis to appear in nature. Gram positive eubacteria, cyanobacteria, Archeae, and eukaryotic organelles possess a noncanonical pathway for the synthesis of Gln-tRNA Gln based on the transformation of Glu-tRNA Gln (synthesized by Glu-tRNA synthetase, GluRS) using the enzyme Glu-tRNA Gln amidotransferase (Glu-AdT). The reactions involved in the transamidation pathway are as follows (Curnow, A. W. et al. (1997) Nucleic Acids Symposium 36:24): A similar enzyme, Asp-tRNA Asn amidotransferase, exists in Archaea, which transforms Asp-tRNA Asn to Asn-tRNA Asn . Formylase, the enzyme that transforms Met-tRNA fMet to fMet-tRNA fMet in eubacteria, is likely to be a related enzyme. A hydrolytic activity has also been identified that destroys mischarged Val-tRNA Ile (Schimmel et al., supra). One likely scenario for the evolution of Glu-AdT in primitive life forms is the absence of a specific glutaminyl-tRNA synthetase (GlnRS), requiring an alternative pathway for the synthesis of Gln-tRNA Gln . In fact, deletion of the Glu-AdT operon in Gram positive bacteria is lethal (Curnow, A. W. et al. (1997) Proc. Natl. Acad. Sci. USA 94:11819-11826). The existence of GluRS activity in other organisms has been inferred by the high degree of conservation in translation machinery in nature; however, GluRS has not been identified in all organisms, including Homo sapiens. Such an enzyme would be responsible for ensuring translational fidelity and reducing the synthesis of defective polypeptides. In addition to their function in protein synthesis, specific aminoacyl tRNA synthetases also play roles in cellular fidelity, RNA splicing, RNA trafficking, apoptosis, and transcriptional and translational regulation. For example, human tyrosyl-tRNA synthetase can be proteolytically cleaved into two fragments with distinct cytokine activities. The carboxy-terminal domain exhibits monocyte and leukocyte chemotaxis activity as well as stimulating production of myeloperoxidase, tumor necrosis factor-α, and tissue factor. The N-terminal domain binds to the interleukin-8 type A receptor and functions as an interleukin-8-like cytokine. Human tyrosyl-tRNA synthetase is secreted from apoptotic tumor cells and may accelerate apoptosis (Wakasugi, K., and P. Schimmel (1999) Science 284:147-151). Mitochondrial Neurospora crassa TyrRS and S. cerevisiae LeuRS are essential factors for certain group I intron splicing activities, and human mitochondrial LeuRS can substitute for the yeast LeuRS in a yeast null strain. Certain bacterial aaRSs are involved in regulating their own transcription or translation (Martini et al., supra). Several aaRSs are able to synthesize diadenosine oligophosphates, a class of signalling molecules with roles in cell proliferation, differentiation, and apoptosis (Kisselev, L. L. et al. (1998) FEBS Lett. 427:157-163; Vartanian, A. et al. (1999) FEBS Lett. 456:175-180). Autoantibodies against aminoacyl-tRNAs are generated by patients with autoimmune diseases such as rheumatic arthritis, dermatomyositis and polymyositis, and correlate strongly with complicating interstitial lung disease (ILD) (Freist, W. et al. (1999) Biol. Chem. 380:623-646; Freist, W. et al. (1996) Biol. Chem. Hoppe Seyler 377:343-356). These antibodies appear to be generated in response to viral infection, and coxsackie virus has been used to induce experimental viral myositis in animals. Comparison of aaRS structures between humans and pathogens has been useful in the design of novel antibiotics (Schimmel et al., supra). Genetically engineered aaRSs have been utilized to allow site-specific incorporation of unnatural amino acids into proteins in vivo (Liu, D. R. et al. (1997) Proc. Natl. Acad. Sci. USA 94:10092-10097). tRNA Modifications The modified ribonucleoside, pseudouridine (Ψ), is present ubiquitously in the anticodon regions of transfer RNAs (tRNAs), large and small ribosomal RNAs (rRNAs), and small nuclear RNAs (snRNAs). Ψ is the most common of the modified nucleosides (i.e., other than G, A, U, and C) present in tRNAs. Only a few yeast tRNAs that are not involved in protein synthesis do not contain Ψ (Cortese, R. et al. (1974) J. Biol. Chem, 249:1103-1108). The enzyme responsible for the conversion of uridine to Ψ, pseudouridine synthase (pseudouridylate synthase), was first isolated from Salmonella typhimurium (Arena, F. et al. (1978) Nucleic Acids Res. 5:4523-4536). The enzyme has since been isolated from a number of mammals, including steer and mice (Green, C. J. et al. (1982) J. Biol. Chen 257:3045-52; and Chen, J. and J. R. Patton (1999) RNA 5:409-419). tRNA pseudouridine synthases have been the most extensively studied members of the family. They require a thiol donor (e.g., cysteine) and a monovalent cation (e.g., ammonia or potassium) for optimal activity. Additional cofactors or high energy molecules (e.g., ATP or GTP) are not required (Green et al., supra). Other eukaryotic pseudouridine synthases have been identified that appear to be specific for rRNA (reviewed in Smith, C. M. and J. A. Steitz (1997) Cell 89:669-672) and a dual-specificity enzyme has been identified that uses both tRNA and rRNA substrates (Wrzesinski, J. et al. (1995) RNA 1: 437-448). The absence of Ψ in the anticodon loop of tRNAs results in reduced growth in both bacteria (Singer, C. E. et al. (1972) Nature New Biol. 238:72-74) and yeast (Lecointe, F. (1998) J. Biol. Chem. 273:1316-1323), although the genetic defect is not lethal. Another ribonucleoside modification that occurs primarily in eukaryotic cells is the conversion of guanosine to N 2 ,N 2 -dimethylguanosine (m 2 2 G) at position 26 or 10 at the base of the D-stem of cytosolic and mitochondrial tRNAs. This posttranscriptional modification is believed to stabilize tRNA structure by preventing the formation of alternative tRNA secondary and tertiary structures. Yeast tRNA Asp is unusual in that it does not contain this modification. The modification does not occur in eubacteria, presumably because the structure of tRNAs in these cells and organelles is sequence constrained and does not require posttranscriptional modification to prevent the formation of alternative structures (Steinberg, S. and R. Cedergren (1995) RNA 1:886-891, and references within). The enzyme responsible for the conversion of guanosine to m 2 2 G is a 63 kDa S-adenosylmethionine (SAM)-dependent tRNA N 2 ,N 2 -dimethyl-guanosine methyltransferase (also referred to as the TRM1 gene product and herein referred to as TRM) (Edqvist, J. (1995) Biochimie 77:54-61). The enzyme localizes to both the nucleus and the mitochondria (Li, J.-M. et al. (1989) J. Cell Biol. 109:1411-1419). Based on studies with TRM from Xenopus laevis, there appears to be a requirement for base pairing at positions C11-G24 and G10-C25 immediately preceding the G26 to be modified, with other structural features of the tRNA also being required for the proper presentation of the G26 substrate (Edqvist. J. et al. (1992) Nucleic Acids Res. 20:6575-6581). Studies in yeast suggest that cells carrying a weak ochre tRNA suppressor (sup3-i) are unable to suppress translation termination in the absence of TRM activity, suggesting a role for TRM in modifying the frequency of suppression in eukaryotic cells (Niederberger, C. et al. (1999) FEBS Lett. 464:67-70), in addition to the more general function of ensuring the proper three-dimensional structures for tRNA. Translation Initiation Initiation of translation can be divided into three stages. The first stage brings an initiator transfer RNA (Met-tRNA f ) together with the 40S ribosomal subunit to form the 43S preinitiation complex. The second stage binds the 43S preinitiation complex to the mRNA, followed by migration of the complex to the correct AUG initiation codon. The third stage brings the 60S ribosomal subunit to the 40S subunit to generate an 80S ribosome at the inititation codon. Regulation of translation primarily involves the first and second stage in the initiation process (Pain, V. M. (1996) Eur. J. Biochem. 236:747-771). Several initiation factors, many of which contain multiple subunits, are involved in bringing an initiator tRNA and the 40S ribosomal subunit together. eIF2, a guanine nucleotide binding protein, recruits the initiator tRNA to the 40S ribosomal subunit. Only when eIF2 is bound to GTP does it associate with the initiator tRNA. eIF2B, a guanine nucleotide exchange protein, is responsible for converting eIF2 from the GDP-bound inactive form to the GTP-bound active form. Two other factors, eIF1A and eIF3 bind and stabilize the 40S subunit by interacting with the 18S ribosomal RNA and specific ribosomal structural proteins. eIF3 is also involved in association of the 40S ribosomal subunit with mRNA. The Met-tRNA f , eFI1A, eIF3, and 40S ribosomal subunit together make up the 43S preinitiation complex (Pain, supra). Additional factors are required for binding of the 43S preinitiation complex to an mRNA molecule, and the process is regulated at several levels. eIF4F is a complex consisting of three proteins: eIF4E, eIF4A, and eIF4G. eIF4E recognizes and binds to the mRNA 5′-terminal m 7 GTP cap, eIF4A is a bidirectional RNA-dependent helicase, and eIF4G is a scaffolding polypeptide. eIF4G has three binding domains. The N-terminal third of eIF4G interacts with eIF4E, the central third interacts with eIF4A, and the C-terminal third interacts with eIF3 bound to the 43S preinitiation complex. Thus, eIF4G acts as a bridge between the 40S ribosomal subunit and the mRNA (Hentze, M. W. (1997) Science 275:500-501). The ability of eIF4F to initiate binding of the 43S preinitiation complex is regulated by structural features of the mRNA. The mRNA molecule has an untranslated region (UTR) between the 5′ cap and the AUG start codon. In some mRNAs this region forms secondary structures that impede binding of the 43S preinitiation complex. The helicase activity of eIF4A is thought to function in removing this secondary structure to facilitate binding of the 43S preinitiation complex (Pain, supra). Translation Elongation Elongation is the process whereby additional amino acids are joined to the initiator methionine to form the complete polypeptide chain. The elongation factors EF1α, EF1βγ, and EF2 are involved in elongating the polypeptide chain following initiation. EF1α is a GTP-binding protein. In EF1α's GTP-bound form, it brings an aminoacyl-tRNA to the ribosome's A site. The amino acid attached to the newly arrived aminoacyl-tRNA forms a peptide bond with the initiatior methionine. The GTP on EF1α is hydrolyzed to GDP, and EF1α-GDP dissociates from the ribosome. EF1βγ binds EF1α-GDP and induces the dissociation of GDP from EF1α, allowing EF1α to bind GTP and a new cycle to begin. As subsequent aminoacyl-tRNAs are brought to the ribosome, EF-G, another GTP-binding protein, catalyzes the translocation of tRNAs from the A site to the P site and finally to the E site of the ribosome. This allows the ribosome and the mRNA to remain attached during translation. Translation Termination The release factor eRF carries out termination of translation. eRF recognizes stop codons in the mRNA, leading to the release of the polypeptide chain from the ribosome. Expression Profiling Microarrays are analytical tools used in bioanalysis. A microarray has a plurality of molecules spatially distributed over, and stably associated with, the surface of a solid support. Microarrays of polypeptides, polynucleotides, and/or antibodies have been developed and find use in a variety of applications, such as gene sequencing, monitoring gene expression, gene mapping, bacterial identification, drug discovery, and combinatorial chemistry. The potential application of gene expression profiling is particularly relevant to improving diagnosis, prognosis, and treatment of disease. One area in particular in which microarrays find use is in gene expression analysis. Array technology can provide a simple way to explore the expression of a single polymorphic gene or the expression profile of a large number of related or unrelated genes. When the expression of a single gene is examined, arrays are employed to detect the expression of a specific gene or its variants. When an expression profile is examined, arrays provide a platform for identifying genes that are tissue specific, are affected by a substance being tested in a toxicology assay, are part of a signaling cascade, carry out housekeeping functions, or are specifically related to a particular genetic predisposition, condition, disease, or disorder. Lung cancer is the leading cause of cancer death in the United States, affecting more than 100,000 men and 50,000 women each year. Nearly 90% of the patients diagnosed with lung cancer are cigarette smokers. Tobacco smoke contains thousands of noxious substances that induce carcinogen metabolizing enzymes and covalent DNA adduct formation in the exposed bronchial epithelium In nearly 80% of patients diagnosed with lung cancer, metastasis has already occurred. Most commonly lung cancers metastasize to pleura, brain, bone, pericardium, and liver. The decision to treat with surgery, radiation therapy, or chemotherapy is made on the basis of tumor histology, response to growth factors or hormones, and sensitivity to inhibitors or drugs. With current treatments, most patients die within one year of diagnosis. Earlier diagnosis and a systematic approach to identification, staging, and treatment of lung cancer could positively affect patient outcome. Lung cancers progress through a series of morphologically distinct stages from hyperplasia to invasive carcinoma. Malignant lung cancers are divided into two groups comprising four histopathological classes. The Non Small Cell Lung Carcinoma (NSCLC) group includes squamous cell carcinomas, adenocarcinomas, and large cell carcinomas and accounts for about 70% of all lung cancer cases. Adenocarcinomas typically arise in the peripheral airways and often form mucin secreting glands. Squamous cell carcinomas typically arise in proximal airways. The histogenesis of squamous cell carcinomas may be related to chronic inflammation and injury to the bronchial epithelium, leading to squamous metaplasia. The Small Cell Lung Carcinoma (SCLC) group accounts for about 20% of lung cancer cases. SCLCs typically arise in proximal airways and exhibit a number of paraneoplastic syndromes including inappropriate production of adrenocorticotropin and anti-diuretic hormone. Lung cancer cells accumulate numerous genetic lesions, many of which are associated with cytologically visible chromosomal aberrations. The high frequency of chromosomal deletions associated with lung cancer may reflect the role of multiple tumor suppressor loci in the etiology of this disease. Deletion of the short arm of chromosome 3 is found in over 90% of cases and represents one of the earliest genetic lesions leading to lung cancer. Deletions at chromosome arms 9p and 17p are also common. Other frequently observed genetic lesions include overexpression of telomerase, activation of oncogenes such as K-ras and c-myc, and inactivation of tumor suppressor genes such as RB, p53 and CDKN2. Genes differentially regulated in lung cancer have been identified by a variety of methods. Using mRNA differential display technology, Manda et al. (1999; Genomics 51:5-14) identified five genes differentially expressed in lung cancer cell lines compared to normal bronchial epithelial cells. Among the known genes, pulmonary surfactant apoprotein A and alpha 2 macroglobulin were down regulated whereas nm23H1 was upregulated. Petersen et al. (2000; Int J. Cancer, 86:512-517) used suppression subtractive hybridization to identify 552 clones differentially expressed in lung tumor derived cell lines, 205 of which represented known genes. Among the known genes, thrombospondin-1, fibronectin, intercellular adhesion molecule 1, and cytokeratins 6 and 18 were previously observed to be differentially expressed in lung cancers. Wang et al. (2000; Oncogene 19:1519-1528) used a combination of microarray analysis and subtractive hybridization to identify 17 genes differentially overexpresssed in squamous cell carcinoma compared with normal lung epithelium Among the known genes they identified were keratin isoform 6, KOC, SPRC, IGFb2, connexin 26, plakofillin 1 and cytokeratin 13. There are more than 180,000 new cases of breast cancer diagnosed each year, and the mortality rate for breast cancer approaches 10% of all deaths in females between the ages of 45-54 (K. Gish (1999) AWIS Magazine 28:7-10). However the survival rate based on early diagnosis of localized breast cancer is extremely high (97%), compared with the advanced stage of the disease in which the tumor has spread beyond the breast (22%). Current procedures for clinical breast examination are lacking in sensitivity and specificity, and efforts are underway to develop comprehensive gene expression profiles for breast cancer that may be used in conjunction with conventional screening methods to improve diagnosis and prognosis of this disease (Perou C. M. et al. (2000) Nature 406:747-752). Breast cancer is a genetic disease commonly caused by mutations in breast epithelial cells. Mutations in two genes, BRCA1 and BRCA2, are known to greatly predispose a woman to breast cancer and may be passed on from parents to children (Gish, supra). However, this type of hereditary breast cancer accounts for only about 5% to 9% of breast cancers, while the vast majority of breast cancer is due to nohinherited mutations that occur in breast epithelial cells. A good deal is already known about the expression of specific genes associated with breast cancer. For example, the relationship between expression of epidermal growth factor (EGF) and its receptor, EGFR, to human mammary carcinoma has been particularly well studied. (See Khazaie, K. et al. (1993) Cancer and Metastasis Rev. 12:255-274), and references cited therein for a review of this area.) Overexpression of EGFR, particularly coupled with down-regulation of the estrogen receptor, is a marker of poor prognosis in breast cancer patients. In addition, EGFR expression in breast tumor metastases is frequently elevated relative to the primary tumor, suggesting that EGFR is involved in tumor progression and metastasis. This is supported by accumulating evidence that EGF has effects on cell functions related to metastatic potential, such as cell motility, chemotaxis, secretion and differentiation. Changes in expression of other members of the erbB receptor family, of which EGFR is one, have also been implicated in breast cancer. The abundance of erbB receptors, such as HER-2/neu, HER-3, and HER-4, and their ligands in breast cancer points to their functional importance in the pathogenesis of the disease, and may therefore provide targets for therapy of the disease (Bacus, S. S. et al. (1994) Am. J. Clin. Pathol. 102:S13-S24). Other known markers of breast cancer include a human secreted frizzled protein mRNA that is downregulated in breast tumors; the matrix G1a protein which is overexpressed is human breast carcinoma cells; Drg1 or RTP, a gene whose expression is diminished in colon, breast, and prostate tumors; maspin, a tumor suppressor gene downregulated in invasive breast carcinomas; and CaN19, a member of the S100 protein family, all of which are down regulated in mammary carcinoma cells relative to normal mammary epithelial cells (Zhou Z. et al. (1998) Int. J. Cancer 78:95-99; Chen, L. et al. (1990) Oncogene 5:1391-1395; Ulrix W. et al (1999) FEBS Lett. 455:23-26; Sager, R. et al. (1996) Curr. Top. Microbiol. Immunol. 213:51-64; and Lee, S. W. et al. (1992) Proc. Natl. Acad. Sci. USA 89:2504-2508). The potential application of gene expression profiling is particularly relevant to measuring the toxic response to potential therapeutic compounds and of the metabolic response to therapeutic agents. Diseases treated with steroids and disorders caused by the metabolic response to treatment with steroids include adenomatosis, cholestasis, cirrhosis, hemangioma, Henoch-Schonlein purpura, hepatitis, hepatocellular and metastatic carcinomas, idiopathic thrombocytopenic purpura, porphyria, sarcoidosis, and Wilson disease. Response may be measured by comparing both the levels and sequences expressed in tissues from subjects exposed to or treated with steroid compounds such as mifepristone, progesterone, beclomethasone, medroxyprogesterone, budesonide, prednisone, dexamethasone, betamethasone, or danazol with the levels and sequences expressed in normal untreated tissue. Steroids are a class of lipid-soluble molecules, including cholesterol, bile acids, vitamin D, and hormones, that share a common four-ring structure based on cyclopentanoperhydrophenanthrene and that carrry out a wide variety of functions. Cholesterol, for example, is a component of cell membranes that controls membrane fluidity. It is also a precursor for bile acids which solubilize lipids and facilitate absorption in the small intestine during digestion. Vitamin D regulates the absorption of calcium in the small intestine and controls the concentration of calcium in plasma. Steroid hormones, produced by the adrenal cortex, ovaries, and testes, include glucocorticoids, mineralocorticoids, androgens, and estrogens. They control various biological processes by binding to intracellular receptors that regulate transcription of specific genes in the nucleus. Glucocorticoids, for example, increase blood glucose concentrations by regulation of gluconeogenesis in the liver, increase blood concentrations of fatty acids by promoting lipolysis in adipose tissues, modulate sensitivity to catcholamines in the central nervous system, and reduce inflammation. The principal mineralocorticoid, aldosterone, is produced by the adrenal cortex and acts on cells of the distal tubules of the kidney to enhance sodium ion reabsorption. Androgens, produced by the interstitial cells of Leydig in the testis, include the male sex hormone testosterone, which triggers changes at puberty, the production of sperm and maintenance of secondary sexual characteristics. Female sex hormones, estrogen and progesterone, are produced by the ovaries and also by the placenta and adrenal cortex of the fetus during pregnancy. Estrogen regulates female reproductive processes and secondary sexual characteristics. Progesterone regulates changes in the endometrium during the menstrual cycle and pregnancy. Steroid hormones are widely used for fertility control and in anti-inflammatory treatments for physical injuries and diseases such as arthritis, asthma, and auto-immune disorders. Progesterone, a naturally occurring progestin, is primarily used to treat amenorrhea, abnormal uterine bleeding, or as a contraceptive. Endogenous progesterone is responsible for inducing secretory activity in the endometrium of the estrogen-primed uterus in preparation for the implantation of a fertilized egg and for the maintenance of pregnancy. It is secreted from the corpus luteum in response to luteinizing hormone (LH). The primary contraceptive effect of exogenous progestins involves the suppression of the midcycle surge of LH. At the cellular level, progestins diffuse freely into target cells and bind to the progesterone receptor. Target cells include the female reproductive tract, the mammary gland, the hypothalamus, and the pituitary. Once bound to the receptor, progestins slow the frequency of release of gonadotropin releasing hormone from the hypothalamus and blunt the pre-ovulatory LH surge, thereby preventing follicular maturation and ovulation. Progesterone has minimal estrogenic and androgenic activity. Progesterone is metabolized hepatically to pregnanediol and conjugated with glucuronic acid. Medroxyprogesterone (MAH), also known as 6α-methyl-17-hydroxyprogesterone, is a synthetic progestin with a pharmacological activity about 15 times greater than progesterone. MAH is used for the treatment of renal and endometrial carcinomas, amenorrhea, abnormal uterine bleeding, and endometriosis associated with hormonal imbalance. MAH has a stimulatory effect on respiratory centers and has been used in cases of low blood oxygenation caused by sleep apnea, chronic obstructive pulmonary disease, or hypercapnia. Mifepristone, also known as RU-486, is an antiprogesterone drug that blocks receptors of progesterone. It counteracts the effects of progesterone, which is needed to sustain pregnancy. Mifepristone induces spontaneous abortion when administered in early pregnancy followed by treatment with the prostaglandin, misoprostol. Further, studies show that mifepristone at a substantially lower dose can be highly effective as a postcoital contraceptive when administered within five days after unprotected intercourse, thus providing women with a “morning-after pill” in case of contraceptive failure or sexual assault. Mifepristone also has potential uses in the treatment of breast and ovarian cancers in cases in which tumors are progesterone-dependent It interferes with steroid-dependent growth of brain meningiomas, and may be useful in treatment of endometriosis where it blocks the estrogen-dependent growth of endometrial tissues. It may also be useful in treatment of uterine fibroid tumors and Cushing's Syndrome. Mifepristone binds to glucocorticoid receptors and interferes with cortisol binding. Mifepristone also may act as an anti-glucocorticoid and be effective for treating conditions where cortisol levels are elevated such as AIDS, anorexia nervosa, ulcers, diabetes, Parkinson's disease, multiple sclerosis, and Alzheimer's disease. Danazol is a synthetic steroid derived from ethinyl testosterone. Danazol indirectly reduces estrogen production by lowering pituitary synthesis of follicle-stimulating hormone and LH. Danazol also binds to sex hormone receptors in target tissues, thereby exhibiting anabolic, antiestrognic, and weakly androgenic activity. Danazol does not possess any progestogenic activity, and does not suppress normal pituitary release of corticotropin or release of cortisol by the adrenal glands. Danazol is used in the treatment of endometriosis to relieve pain and inhibit endometrial cell growth. It is also used to treat fibrocystic breast disease and hereditary angioedema. Corticosteroids are used to relieve inflammation and to suppress the immune response. They inhibit eosinophil, basophil, and airway epithelial cell function by regulation of cytokines that mediate the inflammatory response. They inhibit leukocyte infiltration at the site of inflammation, interfere in the function of mediators of the inflammatory response, and suppress the humoral immune response. Corticosteroids are used to treat allergies, asthma, arthritis, and skin conditions. Beclomethasone is a synthetic glucocorticoid that is used to treat steroid-dependent asthma, to relieve symptoms associated with allergic or nonallergic (vasomotor) rhinitis, or to prevent recurrent nasal polyps following surgical removal. The anti-inflammatory and vasoconstrictive effects of intranasal beclomethasone are 5000 times greater than those produced by hydrocortisone. Budesonide is a corticosteroid used to control symptoms associated with allergic rhinitis or asthma Budesonide has high topical anti-inflammatory activity but low systemic activity. Dexamethasone is a synthetic glucocorticoid used in anti-inflammatory or immunosuppressive compositions. It is also used in inhalants to prevent symptoms of asthma. Due to its greater ability to reach the central nervous system, dexamethasone is usually the treatment of choice to control cerebral edema. Dexamethasone is approximately 20-30 times more potent than hydrocortisone and 5-7 times more potent than prednisone. Prednisone is metabolized in the liver to its active form, prednisolone, a glucocorticoid with anti-inflammatory properties. Prednisone is approximately 4 times more potent than hydrocortisone and the duration of action of prednisone is intermediate between hydrocortisone and dexamethasone. Prednisone is used to treat allograft rejection, asthma, systemic lupus erythematosus, arthritis, ulcerative colitis, and other inflammatory conditions. Betamethasone is a synthetic glucocorticoid with antiinflammatory and immunosuppressive activity and is used to treat psoriasis and fungal infections, such as athlete's foot and ringworm. The anti-inflammatory actions of corticosteroids are thought to involve phospholipase A 2 inhibitory proteins, collectively called lipocortins. Lipocortins, in turn, control the biosynthesis of potent mediators of inflammation such as prostaglandins and leukotrienes by inhibiting the release of the precursor molecule arachidonic acid. Proposed mechanisms of action include decreased IgE synthesis, increased number of β-adrenergic receptors on leukocytes, and decreased arachidonic acid metabolism During an immediate allergic reaction, such as in chronic bronchial asthma, allergens bridge the IgE antibodies on the surface of mast cells, which triggers these cells to release chemotactic substances. Mast cell influx and activation, therefore, is partially responsible for the inflammation and hyperirritability of the oral mucosa in asthmatic patients. This inflammation can be retarded by administration of corticosteroids. The effects upon liver metabolism and hormone clearance mechanisms are important to understand the pharmacodynamics of a drug. The human C3A cell line is a clonal derivative of HepG2/C3 (hepatoma cell line, isolated from a 15-year-old male with liver tumor), which was selected for strong contact inhibition of growth. The use of a clonal population enhances the reproducibility of the cells. C3A cells have many characteristics of primary human hepatocytes in culture: i) expression of insulin receptor and insulin-like growth factor II receptor; ii) secretion of a high ratio of serum albumin compared with α-fetoprotein iii) conversion of ammonia to urea and glutamine; iv) metabolize aromatic amino acids; and v) proliferate in glucose-free and insulin-free medium The C3A cell line is now well established as an in vitro model of the mature human liver (Mickelson et al. (1995) Hepatology 22:866-875; Nagendra et al. (1997) Am J Physiol 272:G408-G416). There is a need in the art for new compositions, including nucleic acids and proteins, for the diagnosis, prevention, and treatment of cell proliferative, DNA repair, neurological, reproductive, developmental, and autoimmune/inflammatory disorders, and infections. |
<SOH> SUMMARY OF THE INVENTION <EOH>Various embodiments of the invention provide purified polypeptides, nucleic acid-associated proteins, referred to collectively as “NAAP” and individually as “NAAP-1,” “NAAP-2,” “NAAP-3,” “NAAP4,” “NAAP-5,” “NAAP-6,” “NAAP-7,” “NAAP-8,” “NAAP-9,” “NAAP-10,” “NAAP-11,” “NAAP-12,” “NAAP-13,” “NAAP-14,” “NAAP-15,” “NAAP-16,” “NAAP-17,” “NAAP-18,” “NAAP-19,” “NAAP-20,” “NAAP-21,” “NAAP-22,” “NAAP-23,” ”NAAP-24,” “NAAP-25,” “NAAP-26,” “NAAP-27,” “NAAP-28,” “NAAP-29,” “NAAP-30,” “NAAP-31,” “NAAP-32,” “NAAP-33,” and methods for using these proteins and their encoding polynucleotides for the detection, diagnosis, and treatment of diseases and medical conditions. Embodiments also provide methods for utilizing the purified nucleic acid-associated proteins and/or their encoding polynucleotides for facilitating the drug discovery process, including determination of efficacy, dosage, toxicity, and pharmacology. Related embodiments provide methods for utilizing the purified nucleic acid-associated proteins and/or their encoding polynucleotides for investigating the pathogenesis of diseases and medical conditions. An embodiment provides an isolated polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. Another embodiment provides an isolated polypeptide comprising an amino acid sequence of SEQ ID NO:1-33. Still another embodiment provides an isolated polynucleotide encoding a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. In another embodiment, the polynucleotide encodes a polypeptide selected from the group consisting of SEQ ID NO:1-33. In an alternative embodiment, the polynucleotide is selected from the group consisting of SEQ ID NO:34-66. Still another embodiment provides a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. Another embodiment provides a cell transformed with the recombinant polynucleotide. Yet another embodiment provides a transgenic organism comprising the recombinant polynucleotide. Another embodiment provides a method for producing a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. The method comprises a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding the polypeptide, and b) recovering the polypeptide so expressed. Yet another embodiment provides an isolated antibody which specifically binds to a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. Still yet another embodiment provides an isolated polynucleotide selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical or at least about 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). In other embodiments, the polynucleotide can comprise at least about 20, 30, 40, 60, 80, or 100 contiguous nucleotides. Yet another embodiment provides a method for detecting a target polynucleotide in a sample, said target polynucleotide being selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical or at least about 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). The method comprises a) hybridizing the sample with a probe comprising at least 20 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide or fragments thereof, and b) detecting the presence or absence of said hybridization complex. In a related embodiment, the method can include detecting the amount of the hybridization complex. In still other embodiments, the probe can comprise at least about 20, 30, 40, 60, 80, or 100 contiguous nucleotides. Still yet another embodiment provides a method for detecting a target polynucleotide in a sample, said target polynucleotide being selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical or at least about 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). The method comprises a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof. In a related embodiment, the method can include detecting the amount of the amplified target polynucleotide or fragment thereof. Another embodiment provides a composition comprising an effective amount of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and a pharmaceutically acceptable excipient. In one embodiment, the composition can comprise an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. Other embodiments provide a method of treating a disease or condition associated with decreased or abnormal expression of functional NAAP, comprising administering to a patient in need of such treatment the composition. Yet another embodiment provides a method for screening a compound for effectiveness as an agonist of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting agonist activity in the sample. Another embodiment provides a composition comprising an agonist compound identified by the method and a pharmaceutically acceptable excipient. Yet another embodiment provides a method of treating a disease or condition associated with decreased expression of functional NAAP, comprising administering to a patient in need of such treatment the composition. Still yet another embodiment provides a method for screening a compound for effectiveness as an antagonist of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting antagonist activity in the sample. Another embodiment provides a composition comprising an antagonist compound identified by the method and a pharmaceutically acceptable excipient. Yet another embodiment provides a method of treating a disease or condition associated with overexpression of functional NAAP, comprising administering to a patient in need of such treatment the composition. Another embodiment provides a method of screening for a compound that specifically binds to a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. The method comprises a) combining the polypeptide with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide to the test compound, thereby identifying a compound that specifically binds to the polypeptide. Yet another embodiment provides a method of screening for a compound that modulates the activity of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical or at least about 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-33. The method comprises a) combining the polypeptide with at least one test compound under conditions permissive for the activity of the polypeptide, b) assessing the activity of the polypeptide in the presence of the test compound, and c) comparing the activity of the polypeptide in the presence of the test compound with the activity of the polypeptide in the absence of the test compound, wherein a change in the activity of the polypeptide in the presence of the test compound is indicative of a compound that modulates the activity of the polypeptide. Still yet another embodiment provides a method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, the method comprising a) exposing a sample comprising the target polynucleotide to a compound, b) detecting altered expression of the target polynucleotide, and c) comparing the expression of the target polynucleotide in the presence of varying amounts of the compound and in the absence of the compound. Another embodiment provides a method for assessing toxicity of a test compound, said method comprising a) treating a biological sample containing nucleic acids with the test compound; b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide selected from the group consisting of i) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, ii) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical or at least about 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, iii) a polynucleotide having a sequence complementary to i), iv) a polynucleotide complementary to the polynucleotide of ii), and v) an RNA equivalent of i)-iv). Hybridization occurs under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide selected from the group consisting of i) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, ii) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical or at least about 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:34-66, iii) a polynucleotide complementary to the polynucleotide of i), iv) a polynucleotide complementary to the polynucleotide of ii), and v) an RNA equivalent of i)-iv). Alternatively, the target polynucleotide can comprise a fragment of a polynucleotide selected from the group consisting of i)-v) above; c) quantifying the amount of hybridization complex; and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound. |
Edible pesticidal formulations |
The invention provides a granular edible pesticidal composition comprising: (a) a continuous hydrophilic matrix phase comprising hydrophilic material, preferably in particulate form and water, said matrix phase being palatable to pests; and (b) a discontinous oleophilic phase dispersed within the hydrophilic matrix phase and comprising an oleophilic carrier and pesticide preferably dissolved in the oleophilic phase. |
1. The invention provides a granular edible pesticidal composition comprising: (a) a continuous hydrophilic matrix phase comprising hydrophilic material, preferably in particulate form and water, said matrix phase being palatable to pests; and (b) a discontinuous oleophilic phase dispersed within the hydrophilic matrix phase and comprising an oleophilic carrier and pesticide preferably dissolved in the oleophilic phase. 2. A granular edible pesticidal composition according to claim 1 wherein the pesticide is dissolved in the oleophilic phase. 3. A granular pesticidal composition according to claim 1 comprising 70 to 95% by weight of hydrophilic matrix phase including water and 5 to 30% of oleophilic phase. 4. A granular pesticidal material according to claim 1 comprising from 0.001 to 33% by weight, based on the weight of oleophilic phase of pesticide. 5. A granular pesticidal composition according to claim 1 wherein the granules have an individual crush strength of at least 500 g. 6. A granular pesticidal composition according to claim 1 comprising from 5 to 20% by weight of the total composition of oleophilic phase and 80 to 95% by weight of the total composition of the hydrophilic matrix phase. 7. A granular pesticidal composition according to claim 1 comprising an oleophilic phase having a Brookfield viscosity at a temperature of 25° C. at least 100 CP. 8. A granular pesticidal composition according to claim 7 wherein the oleophilic phase comprises one or more oleophilic carriers selected from the group consisting of chlorinated hydrocarbons, polyalkylene glycols. 9. A granular pesticidal composition according to claim 7 wherein the oleophilic phase comprises a chlorinated hydrocarbon comprising at least 12 carbon atoms and having a degree of chlorination of at least 40%. 10. A granular pesticidal composition according to claim 1 wherein the hydrophilic material is selected from the group consisting of processes or unprocessed cereal grains, blood and bone, peat and animal manure. 11. A granular pesticidal composition according to claim 1 wherein the hydrophilic material is peat. 12. A granular pesticidal composition according to claim 1 wherein the pesticide has a vapour pressure of at least one millipascal. 13. A granular pesticide according to claim 1 wherein the pesticide is selected from the group consisting of organophosphate insecticides, organochlorine insecticides, carbamate insecticides, synthetic pyrethroids, guanidine/neonicotinoids and mixtures thereof. 14. A granular pesticidal composition according to claim 13 wherein the pesticide is an organophosphate. 15. A granular pesticidal composition according to claim 1 wherein the oleophilic phase provides a drying time test of at least 20%. 16. A granular pesticidal composition according to claim 1 wherein the granules have a maximum dimension in the range of from 0.5 to 10 MM. 17. A granular pesticidal composition according to claim 1 wherein the granules are extruded. 18. A method of controlling ground dwelling pests in a region comprising placing the granular pesticidal composition according claim 1 adjacent or below the surface of the soil 19. A method of preparing a granular pesticidal composition according to claim 1 comprising: (j) mixing water with the hydrophilic phase to form a deformable dough; (ii) spraying an oleophilic phase onto the hydrophilic phase and mixing; (iii) forming the mixture into granules ; and (iv) drying the granules to mechanical integrity. 20. A method according to claim 19 wherein the hydrophilic material comprises fibres and the granules are formed by extrusion. |
<SOH> BACKGROUND <EOH>Edible pesticidal compositions have been widely used in control of pests. In such formulations the loss of active agent is a problem for the efficacy and environmental safety of the composition. The pesticidal agent is often liberated into the environment and is wasted (removed or destroyed) by processes such as volatilization, binding to clay or organic matter, microbial degradation, chemical decay and leaching. This significantly reduces the effective life of the edible pesticidal formulation. Another problem caused by edible pesticidal compositions is that the pesticidal agent is often toxic to beneficial organisms which prey on pests but do not cause feeding damage in their own right. The presence of residual sub-lethal quantities of pesticidal agents through loss of pesticide over time causes pesticide resistance to develop in the population of pests. This problem can be exacerbated by slow release formulations which generate significant zones or “hot spots” of sub-lethal pesticide concentration. Sustained release formulations have been described which provide prolonged pesticidal activity by providing a slow continuous release of pesticide. Such sustained release formulations have been made by containing the pesticidal agent in a hydrophobic matrix material. Example of a controlled release formulation is the SuSCon range of controlled release chlorpyrifos granules sold by Cropcare Australasia Pty Ltd [of 77 Tingira Street, Pinkenba, Queensland, Australia] which are based on the use of thermoplastic resins (such as ethylene-vinyl acetate copolymers) as the matrix phase. Another example of a controlled release formulation is the aphicidal granule product based on the use of thermoplastic resins or wax as described in Australian Patent AU8944301 to ICI PLC. While the slow release of pesticides from these formulations increases the effective life of the edible pesticidal formulation it does not address problems of damage to non-target organisms or the built up of resistance. Many long-lasting hydrophobic matrix materials (e.g. ethylene vinyl acetate copolymers) are not edible by pests and so cannot be used to provide edible pesticidal formulations. Sustained release formulations have also been made by containing the pesticidal agent in a hydrophilic matrix material (i.e. the hydrophilic material provides the continuous phase of the formulation). These hydrophilic materials contain a certain amount of water and may take up more water when they encounter wet conditions. Examples of pesticidal formulations which contain the pesticide in a hydrophilic matrix include: (1) The use of hydrated fibrous mats as carriers by Balassa in U.S. Pat. No. 4,787,928 (2) The use of thermoplastic hydrogels as carriers by Vaughan et al in Australian Patent AU07680991. (3) The use of reversibly dehydrated vegetable and/or fruit to provide rodent baits by Barth et al in EP 86107928 (4) The use of a carrier phase comprising milk solids and sucrose (in the presence of high levels of boric acid as active ingredient) by Nelson et al in U.S. Pat. No. 6,153,181. Nelson points out that toxic baits for crawling insects have historically been water-based, and that water has been presumed necessary for good bait performance. Nelson explains that products comprising significant quantities of water tend to lose effectiveness as a result of water loss, rancidity, break-down of active ingredients etc. (5) The use of an aqueous plant fibre slurry (which is subsequently dried) as the matrix for an agricultural granule has been described by Lowe et al in U.S. Pat. No. 5,019,564. Lowe et al note that the use of clay in the matrix can create chemical inactivation of active ingredients such as chlorpyriphos. (6) The use of polyvinyl alcohol and borate in water (subsequently dried) as a pesticide matrix has been described by Maglio in U.S. Pat. No. RE33,670. (7) The use of portions of corncob in various ratios as a carrier for pesticides has been described by Katz et al in U.S. Pat. No. 4,563,344. The discussion of the background to the invention herein is included to explain the context of the invention. This is not to be taken as an admission that any of the material referred to was published, known or part of the common general knowledge in Australia as at the priority date of any of the claims. None of the above formulations has been shown to provide a long-term ingestible bait which properly contains active ingredient. |
<SOH> SUMMARY OF THE INVENTION <EOH>This invention provides granules comprising: (a) a continuous hydrophilic matrix phase comprising hydrophilic material, preferably in particulate form and water, said matrix phase being palatable to pests; and (b) a discontinuous oleophilic phase dispersed within the hydrophilic matrix phase and comprising an oleophilic carrier and pesticide preferably dissolved in the oleophilic phase. It is surprising that the discontinuously dispersed oleophilic phase enhances the containment of the oil-soluble pesticide because the principal barrier to release would be expected to be the hydrophilic matrix. In one preferment the oleophilic phase is viscous at ambient temperature, i.e. the time taken to pour said oleophilic phase from a 100 ml beaker is in excess of 10 seconds at 20° C. and more preferably in excess of 30 seconds at 20° C. The oleophilic phase will preferably have a Brookfield viscosity greater than 100 cP, more preferably greater than 200 cP. (Measured at a temperature of 25° C.). The invention further provides a method of controlling ground dwelling pests in a region comprising applying the granular pesticidal composition as hereinbefore described adjacent or below the surface of the soil. In regions of thick vegetation the granules may be applied to the thatch of vegetation adjacent the surface of the soil. In yet a further aspect the invention provides a method of preparing a granular pesticidal composition as hereinbefore described comprising: (i) mixing water with the hydrophilic phase to form a deformable dough; (ii) spraying an oleophilic phase onto the hydrophilic phase and mixing; (iii) forming the mixture into granules; and (iv) drying the granules to mechanical integrity. detailed-description description="Detailed Description" end="lead"? |
Beam |
A beam is provided for absorbing impact forces imparted to doors and sides of a vehicle body as a result of collisions from the side as well as from the front and rear of the vehicle. The beam (17, 18, 78, 88) is intended to protect people or goods inside the vehicle against personal injury or damage in the case of such an impact. The beam (17, 18, 78, 88) comprises at least one thin walled, closed profile roll formed member (52, 62, 72, 82) defining a cavity (63, 83). At least one wall of the member defines at least one longitudinally extending groove (53, 65, 73, 81). The beam (17, 18, 78, 88) may also be used as a standard building block for structural components. |
1-27. (canceled) 28. A beam for absorbing impact forces comprising at least one thin walled, closed profile roll formed member of sheet metal defining a cavity wherein at least one wall of the at least one member defines at least one longitudinally extending groove, the at least one groove is orientated substantially perpendicular to the direction of potential impact forces towards the beam, the sheet metal is comprised of a material capable of being hardened and which has a tensile yield limit of approximately 340 MPa in its unhardened state and in its hardened state after accomplished roll forming a tensile yield limit of at least 1100 MPa, the at least one member has at least one predetermined longitudinal section with a yield limit of approximately 340 MPa as compared with a yield point of at least 1100 MPa for the rest of the beam. 29. A beam as claimed in claim 28, wherein the at least one member comprises at least two adjacent members and the adjacent members are connected by an integrally formed web section. 30. A beam as claimed in claim 29, wherein each adjacent member is joined to the integrally formed web section by a seam weld to close the adjacent member. 31. A beam as claimed in claim 28, wherein the at least one longitudinally extending groove is dimensionally sized based upon both wall thickness and lateral length of the at least one wall in which at least one longitudinally extending groove is housed. 32. A beam as claimed in claim 31, wherein the at least one wall has a wall thickness in a range of between 0.7 and 1.0 mm and the at least one longitudinally extending groove has a depth of 3 mm, a width of 3 mm and a small corner radius. 33. A beam as claimed in claim 28, wherein at least one member is non-cylindrical and has corners provided with corner radii which are small relative to the thickness of the sheet metal. 34. A beam as claimed in claim 28, wherein the sheet metal is a steel alloy sheet. 35. A beam as claimed in claim 34, wherein the steel alloy sheet is boron steel. 36. A beam as claimed in claim 28, wherein the sheet metal has a thickness in a range of between 0.7 and 1.8 mm. 37. A beam as claimed in claim 28, wherein the at least one member is closed by a longitudinally extending seam weld. 38. A beam as claimed in claim 37, wherein the seam weld has a yield point of at least 1100 MPa. 39. A beam as claimed in claim 28, wherein at least one wall of the at least one member defines laterally extending stiffening grooves. 40. A beam as claimed in claim 39, wherein the laterally extending grooves project inwardly from the at least one member and are equispaced from one another along the at least one wall of the at least one member. 41. A beam as claimed in claim 28, wherein the at least one longitudinal section with the yield limit of approximately 340 MPa is adapted to be located adjacent a normal sitting position of a car seat occupant. 42. An impact protection assembly for a door having a door frame, an inner door panel and door ends comprising at least an outer beam mounted on the door between the door ends, the outer beam having at least one thin walled, closed profile roll formed member of sheet metal defining a cavity wherein at least one wall of the at least one member defines at least one longitudinally extending groove, the at least one groove is orientated substantially perpendicular to the direction of potential impact forces towards the beam, the sheet metal is comprised of a material capable of being hardened and which has a tensile yield limit of approximately 340 MPa in its unhardened state and in its hardened state after accomplished roll forming a tensile yield limit of at least 1100 MPa, the at least one member has at least one predetermined longitudinal section with a yield limit of approximately 340 MPa as compared with a yield point of at least 1100 MPa for the rest of the beam 43. An impact protection assembly as claimed in claim 42, further comprising an upper beam mounted on a door frame on the inner door panel between the door ends. 44. An impact protection assembly as claimed in claim 43, wherein the upper beam comprises two members connected by a web section and the outer beam comprises a single member. 45. An impact protection assembly as claimed in claim 44, wherein both of the members of the upper beam are provided with a single longitudinally extending groove on one corresponding wall of each member and the at least one member of the outer beam defines two grooves on two opposite walls of the at least one member and laterally extending stiffening grooves on at least one other wall. 46. A method of manufacturing a beam for absorbing impact forces comprising the steps of: (a) providing sheet metal; (b) forming the sheet metal into at least one thin walled, closed profile member having at least one longitudinally extending groove on at least one wall of the member, the at least one groove orientated substantially perpendicular to a direction of potential impact forces towards the beam; (c) locally heating predetermined longitudinal sections of the beam by inductive heating to tempering temperature; and (d) cooling the predetermined longitudinal sections to have independent yield strength values. 47. A method as claimed in claim 46, wherein the sheet metal is a hardenable steel having a tensile yield limit of approximately 340 MPa in an unhardened state and a tensile yield limit of at least 1100 MPa in a hardened state, wherein each predetermined longitudinal section has a yield limit of approximately 340 MPa compared with the yield point of at least 1100 MPa for the rest of the beam. 48. A method as claimed in claim 46, further including the step of roll forming lateral stiffening grooves into at least one wall of the member. 49. A method as claimed in claim 46, wherein the forming step includes the step of roll forming corners on at least one thin walled, closed profile member with corner radii which are small relative to the thickness of the sheet metal. 50. A method as claimed in claim 46, further including the steps of joining a seam with a continuous weld, passing the beam through induction coils to heat the predetermined longitudinal sections to a hardening temperature, spraying water directly onto the beam and cutting the beam to a necessary length. 51. A method as claimed in claim 50, further including the step of applying a tension to a leading edge of the beam to improve the straightness of the beam. 52. A method as claimed in claim 51, wherein the tension is applied by the rollers at the end of the production line positioned to form the beam by bending over the entire length of the beam. 53. A method as claimed in claim 50, wherein the hardening temperature is approximately 920° C. 54. A method as claimed in claim 46, further including the step of applying a tension to a leading edge of the beam to improve the straightness of the beam. 55. A method as claimed in claim 54, wherein the tension is applied by the rollers at the end of the production line positioned to form the beam by bending over the entire length of the beam. 56. A method as claimed in claim 46, further including the step of welding attachment brackets to the at least one thin walled, closed profile member after the cooling step. 57. A method as claimed in claim 46, further including the step of welding the at least one thin walled, closed profile member to adjacent structural members after the cooling step. |
Magnetic bearing spindle |
A magnetic bearing spindle includes a radial magnetic bearing rotor fabricated by laminating a ring-like plate made of electromagnetic steel sheet, an axial magnetic bearing rotor made of magnetic material, and a main shaft motor rotor. A rotation shaft is fit rotatably in these rotors and a plurality of electromagnets are disposed around the rotation shaft. A bracket is provided with a cooling air supply port, and an air passage is provided in a frame. An outer peripheral portion of the axial magnetic bearing rotor is provided with a triangular portion. An outer peripheral portion of an axial direction positioning collar of an axial magnetic bearing stator is provided with through holes. Cooling air is supplied from the through holes. |
1-25. (canceled) 26. A magnetic bearing spindle comprising: a rotation shaft that is held rotatably in a radial magnetic bearing rotor, an axial magnetic bearing rotor, a main shaft motor rotor, and an axial magnetic bearing stator, the axial magnetic bearing stator having an axial direction positioning member, an outer peripheral portion of the axial direction positioning member having a plurality of through holes, the outer peripheral portion of the axial magnetic bearing rotor is formed into a triangular shape or a shape corresponding to a triangular shape; a plurality of electromagnets disposed around the rotation shaft; and a casing that accommodates the rotation shaft and the electromagnets and includes an air passage to pass cooling air from outside to the through holes. 27. The magnetic bearing spindle according to claim 26, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 28. The magnetic bearing spindle according to claim 26, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of helical grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the helical grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 29. The magnetic bearing spindle according to claim 26, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that are substantially parallel to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 30. The magnetic bearing spindle according to claim 26, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that make an angle to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 31. A magnetic bearing spindle comprising: a rotation shaft that is held rotatably in a radial magnetic bearing rotor, an axial magnetic bearing rotor, a main shaft motor rotor, and an axial magnetic bearing stator, the axial magnetic bearing stator having an axial direction positioning member, an outer peripheral portion of the axial direction positioning member having a plurality of first through holes, portions of an outer magnetic pole tooth of the axial magnetic bearing stator, to which the axial magnetic bearing rotor is not opposed and which are not necessary for a magnetic circuit, are provided with a plurality of second through holes that are parallel to an axial direction of the rotation shaft and that communicate with the first through holes; a plurality of electromagnets disposed around the rotation shaft; and a casing that accommodates the rotation shaft and the electromagnets and includes an air passage to pass cooling air from outside to the first through holes. 32. The magnetic bearing spindle according to claim 31, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 33. The magnetic bearing spindle according to claim 31, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of helical grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the helical grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 34. The magnetic bearing spindle according to claim 31, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that are substantially parallel to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 35. The magnetic bearing spindle according to claim 31, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that make an angle to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 36. The magnetic bearing spindle according to claim 31, wherein the outer peripheral portion of the axial magnetic bearing rotor is formed into a triangular shape or a shape corresponding to a triangular shape. 37. A magnetic bearing spindle comprising: a rotation shaft that is held rotatably in a radial magnetic bearing rotor, an axial magnetic bearing rotor, a main shaft motor rotor, an axial magnetic bearing stator, and a main shaft motor stator, the axial magnetic bearing stator having an axial direction positioning member, an outer peripheral portion of the axial direction positioning member having a plurality of first through holes, a plurality of through holes that are parallel to an axial direction of the rotation shaft are provided in a portion of a core back of the radial magnetic bearing rotor and in a first portion of an outer magnetic pole tooth of the axial magnetic bearing stator and in a second portion of a core back of the main shaft motor stator that is disposed in the vicinity of the main shaft motor rotor, the first portion and the second portion being not necessary for a magnetic circuit; a plurality of electromagnets disposed around the rotation shaft; and a casing that accommodates the rotation shaft and the electromagnets and includes an air passage to pass cooling air from outside to the through holes. 38. The magnetic bearing spindle according to claim 37, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 39. The magnetic bearing spindle according to claim 37, wherein an outer peripheral portion of the rotation shaft is provided with a plurality of helical grooves extending in an axial direction of the rotation shaft, the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fitted to a fitting section of the helical grooves such that these rotors are adjacent as well as in contact with each other in the axial direction, thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 40. The magnetic bearing spindle according to claim 37, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that are substantially parallel to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 41. The magnetic bearing spindle according to claim 37, wherein each of the radial magnetic bearing rotor, the axial magnetic bearing rotor, and the main shaft motor rotor are fabricated with a plurality of ring-like plates each having a plurality of notches on an inner periphery by laminating the plates such that the notches form a plurality of grooves that make an angle to the rotation shaft thereby forming an air passage, at least one of outlet and inlet of the air passage is provided with a helical fin which enhances cooling air intake efficiency to the air passage. 42. The magnetic bearing spindle according to claim 37, wherein the outer peripheral portion of the axial magnetic bearing rotor is formed into a triangular shape or a shape corresponding to a triangular shape. |
<SOH> BACKGROUND ART <EOH>Magnetic bearings realize super high speed rotation which is difficult to realize in ball bearings. Magnetic bearings used as machine tool spindles are also aimed at realizing super high speed rotation. Although the magnetic bearing realizes super high speed rotation, the magnetic bearing generates more heat than the ball bearing as the magnetic bearing has more electrical components than the ball bearing. Therefore, a cooing fluid is passed through a rotation shaft to cool the rotation shaft of a spindle unit. For example, Japanese Patent Application Laid-open No. H9-150345, discloses such a cooling apparatus. This cooling apparatus suits well to a spindle unit that realizes a relatively low rotation speed, however, does not suit to a spindle unit that realizes a super high rotation range. Precisely, in the spindle unit that realizes a super high rotation range, mechanical parts that function as a path for the cooling fluid have limitations due to high speed performance, lifetime, and the like. Therefore, if this cooling apparatus used in the spindle unit that realizes a super high rotation range, these mechanical parts get damaged. The present invention has been archived in order to solve the above problem, and it is an object of the present invention to provide a magnetic bearing spindle capable of efficiently cooling the heat source with a simple configuration and capable of being applied to a spindle unit which is used in the super high speed rotation range. |
<SOH> BRIEF DESCRIPTION OF THE DRAWINGS <EOH>FIG. 1 is a sectional view of a magnetic bearing spindle unit according to a first embodiment of the present invention; FIG. 2 ( 1 ) is a front view of an axial positioning collar of an axial magnetic bearing stator, and FIG. 2 ( 2 ) is a sectional view taken along a line A-A in FIG. 2 ( 1 ); FIG. 3 is a sectional view of a magnetic bearing spindle unit according to a second embodiment of the present invention; FIG. 4 ( 1 ) is a front view of an axial magnetic bearing stator, and FIG. 4 ( 2 ) is a sectional view taken along a line B-B in FIG. 4 ( 1 ); FIG. 5 is a sectional view of a magnetic bearing spindle unit according to a third embodiment of the present invention; FIG. 6 is a sectional view of a magnetic bearing spindle unit of a fourth embodiment of the present invention; FIG. 7 is a sectional view of a magnetic bearing spindle unit according to a fifth embodiment of the present invention; FIG. 8 is a perspective view of a rotation shaft shown in FIG. 7 as viewed from a front side of a front radial magnetic bearing rotor; FIG. 9 is a perspective view of the rotation shaft before a radial magnetic bearing rotor, an axial magnetic bearing rotor and a main shaft motor rotor are fitted; FIG. 10 is a sectional view of the rotation shaft which is cut at which each rotor is fitted; FIG. 11 the outer diameter portion of the axial magnetic bearing rotor is formed into a triangular shape or a shape corresponding thereto is a sectional view of a magnetic bearing spindle unit according to a sixth embodiment of the present invention; FIG. 12 is a perspective view of the rotation shaft before the radial magnetic bearing rotor, the axial magnetic bearing rotor and the main shaft motor rotor are fitted; FIG. 13 is a sectional view of a configuration of an essential portion of a magnetic bearing spindle unit according to a seventh embodiment of the present invention; FIG. 14 is a front view of a ring-like plate provided at its inner diameter portion with notches; FIG. 15 is a perspective view of a radial magnetic bearing rotor formed at its inner diameter portion with notched groove extending in the axial direction of the rotor; FIG. 16 is a sectional view of a configuration of an essential portion of a magnetic bearing spindle unit according to an eighth embodiment of the present invention; and FIG. 17 is a perspective view of a radial magnetic bearing rotor formed at its inner diameter portion with helical grooves. detailed-description description="Detailed Description" end="lead"? |
Peptides of melanoma antigen and their use in diagnostic, prophylactic and therapeutic methods |
Immunogenic peptides of a melanoma antigen recognized by T cells, designated gp100, bioassays using the peptides to diagnose, assess or prognose a mammal afflicted with cancer, more specifically melanoma or metastatic melanoma, and use of the proteins and peptides as immunogens to inhibit, prevent or treat melanoma. |
1. An immunogenic peptide having a gp100 amino acid sequence of 40 contiguous amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, said peptide being reactive with T cells or inducing an immune response. 2. The immunogenic peptide of claim 1, wherein such peptide or variant thereof has an amino acid sequence comprising 17 to 21 amino acids in length. 3. The immunogenic peptide of claim 1, wherein said peptide has a sequence TGRAMLGTHTMEVTVYH (SEQ ID NO:2), LSIGTGRAMLGTHTMEVTVYH (SEQ ID NO:3), IGTGRAMLGTHTMEVTVYHRR (SEQ ID NO:4), TGRAMLGTHTMEVTVYHRRGS (SEQ ID NO:5), TGRAMLGTHTMEVTVYHR (SEQ ID NO:6), TGRAMLGTHTMEVTVYHRR (SEQ ID NO:7), TGRAMLGTHTMEVTVYHRRG (SEQ ID NO:8), GTGRAMLGTHTMEVTVYHRRG (SEQ ID NO:9), GTGRAMLGTHTMEVTVYH (SEQ ID NO:10), IGTGRAMLGTHTMEVTVYH (SEQ ID NO:11), SIGTGRAMLGTHTMEVTVYH (SEQ ID NO:12), SIGTGRAMLGTHTMEVTVYHR (SEQ ID NO:13), PVSGLSIGTGRAMLGTHTMEV (SEQ ID NO:20), TTEWVETTARELPIPE (SEQ ID NO: 21), or a variant of any of the foregoing. 4. The immunogenic peptide of claim 1, wherein said peptide has a sequence SGLSIGTGRAMLGTHTMEVTV (SEQ ID NO:14), RAMLGTHTMEVTVYHRRGSRS (SEQ ID NO:15), MLGTHTMEVTVYHRRGSRSYV (SEQ ID NO:16), or variant thereof. 5. The immunogenic peptide of claim 1, wherein the peptide has a sequence TGRAFLGTHTMEVTVYHRRGS (SEQ ID NO:17), TGRALLGTHTMEVTVYHRRGS (SEQ ID NO:18), or TGRAYLGTHTMEVTVYHRRGS (SEQ ID NO:19). 6. The immunogenic peptide of claim 1, wherein said peptide is recognized by HLA-DRβ1*0701 restricted T cells. 7. The immunogenic peptide of claim 1, wherein said peptide is selected from the group consisting of a native, a synthetic or a recombinant peptide. 8. A pharmaceutical composition comprising an immunogenic peptide and a pharmaceutically-acceptable carrier therefor, said peptide having a contiguous gp100 amino acid sequence of 40 amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, said peptide being reactive with T cells. 9. The pharmaceutical composition of claim 8, wherein said immunogenic peptide is selected from any one of SEQ ID NOs:2-21. 10. A method of transducing antigen presenting cells with a gp100 nucleic acid sequence encoding an amino acid sequence of SEQ ID NOs:1-21 comprising: a) obtaining T cells; b) introducing the gp100 nucleic acid sequence into the antigen presenting cells; and c) stimulating the T cells. 11. A method of determining immunogenicity of a peptide having a contiguous gp100 amino acid sequence of 40 amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, said peptide being capable of reacting with T cells, comprising: a) incubating the peptide with mammalian cells; b) exposing the mammalian cells incubated with the immunogenic peptide to T cells; and c) screening for T cell recognition of the peptide using the mammalian cells incubated with the immunogenic peptide, whereupon the immunogenicity of the peptide is determined. 12. The method of claim 11, wherein the immunogenic peptide is selected from any of SEQ ID NOs:2-21. 13. A method of detecting a gp100 immunogenic peptide in a biological sample, wherein said peptide has a contiguous gp100 amino acid sequence of 40 amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, comprising: a) contacting a reagent which specifically reacts and forms a complex with the gp100 peptide in said sample; and b) detecting the formation of the complex between the peptide and the reagent, whereupon the gp100 immunogenic peptide in the biological sample is detected. 14. The method of claim 13, wherein said immunogenic peptide is selected from any one of SEQ ID NOs:2-21. 15. A method of treating melanoma in a mammal comprising administering an effective amount of an immunogenic peptide having a contiguous gp100 amino acid sequence of 40 amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, effective to stimulate an immunotherapeutic response to the melanoma, whereupon the mammal is treated for melanoma. 16. The method of claim 15, wherein the immunogenic peptide is selected from any one of SEQ ID NOs:2-21. 17. A method of preventing or inhibiting a recurrence of melanoma in a mammal comprising administering an effective amount of an immunogenic peptide having a contiguous gp100 amino acid sequence of 40 amino acids or less comprising MLGTHTMEVTV (SEQ ID NO:1) or a variant thereof, effective to stimulate an immunoprophylactic response to the melanoma, whereupon the recurrence of the melanoma is prevented or inhibited. 18. The method of claim 17, wherein the immunogenic peptide is selected from any one of SEQ ID NOs:2-21. 19. The method of claim 13, wherein the sample is selected from the group consisting of mammalian tissues, mammalian cells, necropsy samples, pathology samples and biopsy samples. 20. The method of claim 13, wherein the biological sample is from a mammal afflicted with a disease. 21. The method of claim 20, wherein the biological sample is from a mammal afflicted with melanoma or metastatic melanoma. 22. The method of claim 13, wherein the reagent is an antibody or fragment thereof. 23. The method of claim 22, wherein the reagent is a monoclonal antibody. 24. The method of claim 22, wherein the reagent is a polyclonal antibody. 25. The method of claim 13, wherein the detection of the complex is used to diagnose, assess or prognose a disease state. |
<SOH> BACKGROUND OF THE INVENTION <EOH>Melanomas are aggressive, frequently metastatic tumors derived from either melanocytes or melanocyte related nevus cells (“Cellular and Molecular Immunology” (1991) (eds) Abbas A. K., Lechtman, A. H., Pober, J. S.; W. B. Saunders Company, Philadelphia: pages 340-341). Melanomas make up approximately three percent of all skin cancers and the worldwide increase in melanoma is unsurpassed by any other neoplasm, with the exception of lung cancer in women (“Cellular and Molecular Immunology” (1991) (eds) Abbas, A. K., Lechtiman, A. H., Pober, J. S.; W. B. Saunders Company Philadelphia pages: 340-342;. Kirkwood and Agarwala, (1993) Principles and Practice of Oncology 7:1-16). Even when melanoma is apparently localized to the skin, up to 30% of the patients will develop systemic metastasis and the majority will die (Kirkwood and Agarwala, (1993) Principles and Practice of Oncology 7:1-16). Classic modalities of treating melanoma include surgery, radiation and chemotherapy. In the past decade immunotherapy and gene therapy have emerged as new and promising methods for treating melanoma. T cells play an important role in tumor regression in most murine tumor models. Tumor infiltrating lymphocytes (TIL) that recognize unique cancer antigens can be isolated from many murine tumors. The adoptive transfer of these TIL plus interleukin-2 can mediate the regression of established lung and liver metastases (Rosenberg, S. A., et al., (1986) Science 233:1318-1321). In addition, the secretion of IFN-γ by injected TIL significantly correlates with in vivo regression of murine tumors, suggesting activation of T cells by the tumor antigens (Barth, R. J., et al., (1991) J. Exp. Med. 173:647-658). The known ability of tumor TIL to mediate the regression of metastatic cancer in 35 to 40% of melanoma patients when adoptively transferred into patients with metastatic melanoma attests to the clinical importance of the antigens recognized (Rosenberg, S. A., et al., (1988) N. Enql. J. Med. 319:1676-1680; Rosenberg, S. A., (1992) J. Clin. Oncol. 10:180-199). T cell receptors on CD8 + T cells recognize a complex consisting of an antigenic peptide (9-10 amino acids for HLA-A2), β-2 microglobulin and class I major histocompatibility complex (MHC) heavy chain (HLA-A, B, C, in humans). Peptides generated by digestion of endogenously synthesized proteins are transported into the endoplastic reticulum, bound to class I MHC heavy chain and β2 microglobulin, and finally expressed in the cell surface in the groove of the class I MHC molecule. Thus, T cells can detect molecules that originate from proteins inside cells, in contrast to antibodies that detect intact molecules expressed on the cell surface. Therefore, antigens recognized by T cells may be more useful than antigens recognized by antibodies. Although emphasis is on CD8+ T cell responses, there is emerging support that CD4+ T cells may play an important role in anti-tumor immunity. As reviewed by Pardoll and Topalian ( Curr. Opin. Immunol. 10:588, 1998), CD4+ T cells have been demonstrated in murine studies to exert helper activity through the induction of CD8+ T cells and B cells and further have both direct and indirect effects on tumor cells, including those deficient in MHC class II. In humans, CD4+ T cells play a critical role in the initiation of several autoimmune diseases (Parry et al., Curr. Opin. Immunol. 10:663, 1998) and in pathogenic resistance (Mata and Paterson, J. Immunol. 163:1449, 1999; Zajac et al., J. Exp. Med. 188:2205, 1998). CD4+ T cells activated dendritic cells primarily through the interaction of CD40 and its ligand. There is growing support that the combination of MHC class I and class II epitopes derived from the same tumor antigen can enhance antitumor effector function and long-term immunity (Surman et al., J. Immunol. 164:562, 2000; Ossendorp et al., J. Exp. Med. 187:693, 1998; Matloubian et al., J. Virol. 68:8056, 1994). Strong evidence that an immune response to cancer exists in humans is provided by the existence of lymphocytes within melanoma deposits. These lymphocytes, when isolated, are capable of recognizing specific tumor antigens on autologous and allogeneic melanomas in an MHC-restricted fashion (Itoh, K. et al. (1986), Cancer Res. 46: 3011-3017; Muul, L. M., et al. (1987), J. Immunol. 138:989-995); Topalian, S. L., et al., (1989) J. Immunol. 142: 3714-3725; Darrow, T. L., et al., (1989) J. Immunol. 142: 3329-3335; Hom, S. S., et al., (1991) J. Immunother. 10:153-164; Kawakami, Y., et al., (1992) J. Immunol. 148: 638-643; Hom, S. S., et al., (1993) J. Immunother. 13:18-30; and O'Neil, B. H., et al., (1993) J. Immunol. 151: 1410-1418). TIL from patients with metastatic melanoma recognize shared antigens including melanocyte-melanoma lineage specific tissue antigens in vitro (Kawakami, Y., et al., (1993) J. Immunother. 14: 88-93; Anichini, A. et al., (1993) et al., J. Exp. Med. 177: 989-998). Anti-melanoma T cells appear to be enriched in TIL, probably as a consequence of clonal expansion and accumulation at the tumor site in vivo (Sensi, M., et al., (1993) J. Exp. Med. 178:1231-1246). The fact that many melanoma patients mount cellular and humoral responses against these tumors and that melanomas express both MHC antigens and tumor associated antigens (TAA) suggests that identification and characterization of additional melanoma antigens will be important for immunotherapy of patients with melanoma. The melanocyte differentiation antigen, gp100, is expressed in more than 75% of human melanomas (Cormier et al., Int. J. Cancer 75:517, 1998). Although the gp100 antigen is predominantly expressed intracellularly, it is a suitable immunogenic antigen. The intracellular proteins have been demonstrated to be processed and presented as peptides in the context of MHC molecules to immune system cells. In particular, TIL derived from tumors of melanoma patients have been identified and react with the gp100 antigen. Given that vaccination with a modified gp100 CD8+ T cell epitope combined with IL-2 reportedly resulted in a 42% response rate in patients with metastatic melanoma (Rosenberg et al., Nat. Med. 4:321, 1998; Parkhurst et al., J. Immunol. 157:2539, 1996), only a few patients responded clinically to this particular vaccine regimen, and additionally, only transient responses were observed. Thus, in order to increase the immunogenicity and therapeutic efficacy of vaccines comprising gp100 CD8+ T cell epitopes, antigen-specific CD4+ T cells can be combined. Therefore, the gp100 MHC class I and class II epitopes can be useful for cellular responses against melanoma, and can also play a significant role in therapy and diagnosis of melanoma patients. Peripheral blood lymphocytes have been used to identify several potential melanoma tumor antigens. For example, Van Der Bruggen et al. ( Science 254: 1643-1647, 1991) has characterized a gene coding for a melanoma antigen, designated MAGE-1, using T cell clones established from the peripheral blood of patients who were repetitively immunized in vivo with mutagenized tumor cells. Cytotoxic T cells derived from the peripheral blood lymphocytes of patients with melanoma were used to identify a potential antigenic peptide encoding MAGE-1 (Traversari, C., et al. (1992) J. Exp. Med. 176:1453-1457). Brichard et al. ((1993) J. Exp. Med. 178:489-495) has also characterized a gene encoding a melanoma antigen designated tyrosinase using peripheral blood lymphocytes from patients who were sensitized by repetitive in vitro stimulation with tumor. Further support for the therapeutic potential of melanoma antigens is provided by Brown et al. (U.S. Pat. No. 5,262,177). Brown et al. (U.S. Pat. No. 5,262,177) relates to a recombinant vaccinia virus-based melanoma vaccine where the melanoma antigen p97 is reported to show a protective effect from tumor cell challenge in a murine model. Characterization of additional melanoma antigens can be important for the development of new strategies for cancer immunotherapy, in particular for melanoma. |
<SOH> SUMMARY OF THE INVENTION <EOH>This invention relates, in general, to peptides or variations of peptides derived from a melanoma-associated antigen known as gp100. In addition, the invention relates to methods of using the gp100 peptides or derivatives thereof for treating and preventing the progression of melanoma-associated diseases. Further, the gp100 peptides and variants thereof can be used as an immunogen for the treatment of patients in need thereof. The present invention provides immunogenic peptides derived from a gp100 melanoma antigen protein sequence. For example, new peptides are exemplified by SEQ ID NOs:2-21. In addition, the present invention provides compositions or immunogens comprising all or part of the gp100 protein or peptides capable of eliciting an immune response in a mammal to melanoma antigens. Further provided are diagnostic methods for human disease involving gp100 peptides or variants thereof, in particular for melanomas and metastatic melanomas. Still further provided are therapeutic methods for the treatment of melanoma using all or part of the gp100 peptides, variants, vaccines or immunogens thereof. Yet still further provided are methods of inhibiting or preventing the recurrence of melanoma in a subject in need thereof. |
Heat stable filter construction |
A filter construction includes a pair of pleated filter elements (31, 32) mounted back to back to form a fluted double filter element and having their edges sealed by a sealant or being folded over within U-sectioned resistant edge strips The filter elements comprise a foraminous metal member (12) embedded within or faced on each side with a layer of non-women material (10), which is hydro entangled through the apertures of the metal member (12) and may include stainless steel fibres and the surface laminated or coated with a fluoropolymer. |
1. A filter construction comprising a filter medium, the filter medium comprising a composite structure of a foraminous metal reinforcement having opposing side faces and a consolidated non-woven fibrous material on one or more faces of the metal reinforcement, wherein the fibrous material is consolidated by hydro entanglement jets, the jets passing through apertures of the foraminous metal reinforcement. 2. (Canceled) 3. A filter construction according to claim 1, wherein the metal reinforcement comprises a sheet of metal formed with a multitude of apertures. 4. A filter construction according to claim 1, wherein the metal reinforcement comprises a fine or ultra-fine wire mesh. 5. A filter construction according to claim 1, wherein the non-woven fibrous material includes metal fibres blended with polymer fibres or with a blend of polymer fibres. 6. A filter construction according to claim 1, wherein the non-woven fibrous material comprises high temperature resistant fibres with a porous fluoropolymer coating. 7. A filter construction according to claim 6, wherein the porous fluoropolymer coating contains carbon powder. 8. A filter construction according to claim 1, wherein the filter medium is formed as two pleated sheets which are disposed back to back to form a fluted double filter element, the edges of the pleated sheets being joined by non-adhesive means. 9. A filter construction according to claim 8 wherein the edges of the pleated sheets are joined by folding over overlapping edges of the sheets, and secured by U-sectioned resistant metal side strips. 10. A filter construction according to claim 8 wherein inserts are placed in the re entrant pleats of the sheets, or over the outward ridges thereof to reinforce the filter medium sheets. 11. A filter construction according to claim 8, wherein the overlapping edges of the pleated sheets are secured by a high temperature sealant. |
Engineered rna translocators |
The present invention provides methods and compositions useful for transporting nucleic acids between cells in plants. |
1. An expression cassette comprising a polynucleotide linked to a heterologous nucleic acid, wherein: the polynucleotide comprises an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23, the polynucleotide does not include a nucleotide sequence encoding an active PVX replicase, movement protein or coat protein; and introduction of the expression cassette into a plant tissue expressing TGBp 1-3 and a PVX coat protein results in transport between cells of an RNA molecule comprising ERT. 2. The expression cassette of claim 1, wherein the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:1. 3. The expression cassette of claim 1, wherein the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:5. 4. The expression cassette of claim 3, wherein the ERT sequence is SEQ ID NO:5. 5. The expression cassette of claim 1, wherein the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:1. 6. The expression cassette of claim 5, wherein the ERT sequence is SEQ ID NO:1. 7. The expression cassette of claim 5, wherein the ERT sequence comprises SEQ ID NO:2. 8. The expression cassette of claim 5, wherein the ERT sequence comprises SEQ ID NO:3. 9. The expression cassette of claim 5, wherein the ERT sequence comprises SEQ ID NO:4. 10. The expression cassette of claim 1, further comprising a promoter operably linked to the polynucleotide. 11. The expression cassette of claim 10, wherein the promoter is constitutive. 12. The expression cassette of claim 10, wherein the promoter is inducible or tissue-specific. 13. A cell comprising: (a) an RNA molecule comprising a polynucleotide linked to a heterologous nucleic acid, wherein the polynucleotide comprises an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23; (b) PVX movement proteins TGBp1-3; and (c) a PVX coat protein. 14. The cell of claim 13, wherein the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:1. 15. The cell of claim 13, wherein the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:5. 16. The cell of claim 13, which is a plant cell. 17. The cell of claim 16, wherein the plant cell is part of a plant. 18. The cell of claim 14, wherein the ERT sequence comprises SEQ ID NO:1. 19. The cell of claim 14, wherein the ERT sequence comprises SEQ ID NO:2. 20. The cell of claim 14, wherein the ERT sequence comprises SEQ ID NO:3. 21. The cell of claim 14, wherein the ERT sequence comprises SEQ ID NO:4. 22. A method of mobilizing RNA molecules between cells in a plant, the method comprising, expressing an RNA molecule in a plant cell, the RNA molecule comprising an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23, wherein the ERT sequence is linked to a heterologous polynucleotide. 23 The method of claim 22, the method further comprising expressing PVX movement proteins TGBp1-3 and a PVX coat protein in the plant cell. 24. The method of claim 22, wherein the plant cell expressing the RNA molecule is contained in a plant tissue that is grafted onto the plant. 25. The method of claim 24, wherein the plant is not transgenic. 26. The method of claim 22, wherein the PVX movement and coat proteins are expressed from a viral vector. 27. The method of claim 22, wherein the PVX movement and coat proteins are encoded by a polynucleotide integrated into the plant genome. 28. The method of claim 22, wherein the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:1. 29. The method of claim 22, wherein the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:5. 30. The method of claim 28, wherein the ERT sequence comprises SEQ ID NO:1. 31. The method of claim 29, wherein the ERT sequence comprises SEQ ID NO:5. 32. The method of claim 28, wherein the ERT sequence comprises SEQ ID NO:2. 33. The method of claim 28, wherein the ERT sequence comprises SEQ ID NO:3. 34. The method of claim 28, wherein the ERT sequence comprises SEQ ID NO:4. 35. The method of claim 22, wherein the TGBp1-3 and the PVX coat protein are expressed from a viral genome. 36. The method of claim 22, wherein the TGBp1-3 and the PVX coat protein are expressed from an integrated transgene. 37. A method of identifying a nucleic acid sequence that is transported between cells in a plant, the method comprising, providing at least one polynucleotide comprising a nucleic acid sequence linked to a reporter gene; introducing the polynucleotide into a target plant cell in the plant; and determining whether the reporter gene is expressed in plant cells in the plant other than the target plant cell, thereby identifying a nucleic acid sequence that is transported between cells in a plant. 38. The method of claim 37, wherein the reporter gene is selected from green fluorescence protein, luciferase and P-glucuronidase. 39. The method of claim 37, wherein the nucleic acid sequence is from a plant. 40. A method of mobilizing RNA molecules between cells in a plant, the method comprising, expressing an RNA molecule in a plant cell, the RNA molecule comprising the nucleic acid sequence identified in claim 37 linked to a heterologous polynucleotide. |
<SOH> BACKGROUND OF THE INVENTION <EOH>Asymmetric distribution of mRNA within cells is controlled by protein-RNA interaction. These nucleotide-specific cis-acting elements, or “zip codes” (Bassell, et al., FASEB J. 13:447-454 (1999)), potentiate subcellular delivery and localized protein synthesis (Jansen, R. P. FASEB J. 13, 455-466 (1999)). This process underlies a wide range of cellular and developmental events (Bassell et al, supra, Choi, S. B. et al. Nature 407:765-767 (2000); Roegiers, et al., Trends Cell Biol 10:220-224 (2000)). A new twist to RNA function is the emerging paradigm of its involvement in non-cell-autonomous control of aberrant RNA and virus/transposon challenge (Mourrain, P. et al. Cell 101:533-542 (2000); Dalmay, et al., Cell 101:543-553 (2000); Tabara, H. et al., Cell 99:123-32 (1999); Bosher, et al., Nature Cell Biol. 2:E31-6 (2000)). Here, mobile nucleotide-specific molecules mediate targeted RNA degradation, via RNA interference (RNAi) in animals (Fire, A. et al., Nature 391:806-811 (1998); Fire, A., Trends Genet. 15:358-363 (1999)) or post transcriptional gene silencing (PTGS) in plants (Napoli, et al., Plant Cell 2:279-289 (1990); Fagard, et al., Annu. Rev. Plant Physiol. Plant Mol. Biol. 51:167-194 (2000)). The plant vascular system, and specifically the phloem, serves as the conduit for this selective movement of nucleic acids to distant organs (Palauqui, et al. EMBO J. 16:4738-4745 (1997); Ruiz-Medrano, et al. Development 126:4405-4419 (1999); Xoconostle-Cázares, et al. Science 283:94-98 (1999)). Analysis of plant virus movement has provided insights into the trafficking of proteins and nucleic acid complexes within the body of the plant (Fujiwara, et al. Plant Cell 5:1783-1794 (1993); Noueiry, et al. Cell 76:925-932 (1994)). Supracellular control of plant developmental and physiological processes is mediated at the molecular level by hormones, small molecules, proteins and ribonucleoprotein complexes (RNPs). See, e.g., Lucas, et al. Curr. Opin. Cell Biol. 7:673-680 (1995). Cell-to-cell trafficking of transcription factors, such as KNOTTED-1 (Lucas, et al. Science 270:1980-1983 (1995)) and LEAFY (Sessions, Science 289:779-782 (2000)), via plasmodesmata, demonstrated that plants have evolved a unique mechanism to control cell fate within meristematic tissues. In addition, it has recently been established that mRNA synthesised in one organ is translocated, via the phloem, to distantly-located tissues and organs. This process involves the selective entry and exit of mRNA through the plasmodesmata which interconnect the specialised companion cells to the enucleate sieve elements that comprise the conduit for long-distance transport of nutrients and information molecules. Little is currently known concerning the mechanisms that underlie the selective trafficking of proteins and RNPs through plasmodesmata. The present invention addresses this and other problems. |
<SOH> BRIEF SUMMARY OF THE INVENTION <EOH>This invention provides polynucleotide sequences comprising engineered translocator (ERT) sequences. In some embodiments, the invention provide expression cassettes comprising a polynucleotide linked to a heterologous nucleic acid, wherein: the polynucleotide comprises an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23, the polynucleotide does not include a nucleotide sequence encoding an active PVX replicase, movement protein or coat protein; and introduction of the expression cassette into a plant tissue expressing TGBp1-3 and a PVX coat protein results in transport between cells of an RNA molecule comprising ERT. In some embodiments, the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:1. In some embodiments, the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:5. In some embodiments, the ERT sequence is SEQ ID NO:5. In some embodiments, the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:1. In some embodiments, the ERT sequence is SEQ ID NO:1. In some embodiments, the ERT sequence comprises SEQ ID NO:2. In some embodiments, the ERT sequence comprises SEQ ID NO:3. In some embodiments; the ERT sequence comprises SEQ ID NO:4. In some embodiments, the expression cassette further comprises a promoter operably linked to the polynucleotide. In some embodiments, the promoter is constitutive. In some embodiments, the promoter is inducible or tissue-specific. The present invention also provides cells comprising: (a) an RNA molecule comprising a polynucleotide linked to a heterologous nucleic acid, wherein the polynucleotide comprises an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23; (b) PVX movement proteins TGBp1-3; and (c) a PVX coat protein. In some embodiments, the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:1. In some embodiments, the polynucleotide comprises an ERT sequence at least 70% identical to SEQ ID NO:5. In some embodiments, the cell is a plant cell. In some embodiments, the plant cell is part of a plant. In some embodiments, the ERT sequence comprises SEQ ID NO:1. In some embodiments, the ERT sequence comprises SEQ ID NO:2. In some embodiments, the ERT sequence comprises SEQ ID NO:3. In some embodiments, the ERT sequence comprises SEQ ID NO:4. The invention also provides methods of mobilizing RNA molecules between cells in a plant. In some embodiments, the methods comprise expressing an RNA molecule in a plant cell, the RNA molecule comprising an ERT sequence at least 70% identical to a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:22 and SEQ ID NO:23, wherein the ERT sequence is linked to a heterologous polynucleotide. In some embodiments, the methods further comprise expressing PVX movement proteins TGBp1-3 and a PVX coat protein in the plant cell. In some embodiments, the plant cell expressing the RNA molecule is contained in a plant tissue that is grafted onto the plant. In some embodiments, the plant is not transgenic. In some embodiments, the PVX movement and coat proteins are expressed from a viral vector. In some embodiments, the PVX movement and coat proteins are encoded by a polynucleotide integrated into the plant genome. In some embodiments, the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:1. In some embodiments, the ERT sequence comprises a polynucleotide at least 70% identical to SEQ ID NO:5. In some embodiments, the ERT sequence comprises SEQ ID NO:1. In some embodiments, the ERT sequence comprises SEQ ID NO:5. In some embodiments, the ERT sequence comprises SEQ ID NO:2. In some embodiments, the ERT sequence comprises SEQ ID NO:3. In some embodiments, the ERT sequence comprises SEQ ID NO:4. In some embodiments, the TGBp1-3 and the PVX coat protein are expressed from a viral genome. In some embodiments, the TGBp 1-3 and the PVX coat protein are expressed from an integrated transgene. The invention also provides methods of identifying a nucleic acid sequence that is transported between cells in a plant. In some embodiments, the methods comprise providing at least one polynucleotide comprising a nucleic acid sequence linked to a reporter gene; introducing the polynucleotide into a target plant cell in the plant; and determining whether the reporter gene is expressed in plant cells in the plant other than the target plant cell, thereby identifying a nucleic acid sequence that is transported between cells in a plant. In some embodiments, the reporter gene is selected from green fluorescence protein, luciferase and β-glucuronidase. In some embodiments, the nucleic acid sequence is from a plant. The invention also provides methods of mobilizing RNA molecules between cells in a plant. In some embodiments, the method comprises expressing an RNA molecule in a plant cell, the RNA molecule comprising the nucleic acid sequence identified in claim 37 linked to a heterologous polynucleotide. |
Device and method for controlling the temperature of preforms |
A device and a method used to temper preforms in the manufacture of bottles, in which a tempering device is provided to selectively and directly temper the preforms, and is inserted into preforms. The method provides that the preforms are directly tempered from the inside. |
1. Device (7, 8) to temper preforms (2) in the manufacture of bottles, comprising a tempering device (1) to selectively and directly temper preforms (2), the tempering device (1) adapted to be inserted in the preforms (2). 2. Device according to claim 1, wherein the tempering device (1) is adapted so that the preforms (2) can be tempered in zones (10) that are one of along, perpendicular to, and at an angle to the preform axis. 3. Device according to claim 1, and a preheating device (6) is provided to evenly heat the preforms (2). 4. Device according to claim 1, wherein the tempering device (1) is a radiant heater to selectively heat the preforms (2). 5. Device according to claim 1, wherein the tempering device (1) comprises at least one gas conducting device and at least one gas outlet (3) for contacting the preforms (2) with one of cooled and heated gas. 6. Device according to claim 5, wherein the tempering device (1) comprises at least one gas inlet to receive gas from the preforms. 7. Device according to claim 5, wherein one of the gas inlet (3) and the gas outlet is formed by one of a slot (3), a hole (3), a nozzle (3) and other aperture (3). 8. Device according to claim 5, wherein several gas inlets (2) and several gas outlets are arranged in one of circles and lines. 9. Device according to claim 5, wherein the tempering device comprises a first (1a) and a second tubular element (2b) that move in relationship to each other, whereby the gas outlet arrangement of the first tubular element (1a) is in a first, relative position of the tubular elements that can move in relationship to each other, whereby the first relative position causes an overlap with the gas outlet arrangement of the second tubular element (1b) to provide selective tempering, and the gas outlet arrangement of the first tubular elements (1a) is in a second relative position of the tubular elements that can move in relationship to each other, whereby the second relative position causes an overlap with the gas outlet arrangement of the second tubular element (1b) to provide even tempering. 10. Device according to claim 1, wherein the tempering device (1) is at least partially integrated in the stretch rod of a blow molding device (7). 11. Device according to claim 3, wherein the tempering device (1) is provided on a transport device (8) by means of which the preheated preforms (2) released by the preheating device (6) can be transported to a blow mold device (9). 12. Device according to claim 1, and means to move the tempering device (1) in and out of the preform. 13. Method for tempering preforms (2) used in the manufacture of bottles, comprising the steps of selectively interactively tempering the preforms (2) and directly tempering the inside of the preforms. 14. Method according to claim 13, and the step of selectively tempering the preforms in zones (10) which are one of along, perpendicular to, and at an angle to the preform axis. 15. Method according to claim 13, and the step of evenly heating the preforms (2) before the step of selective tempering. 16. Method according claim 13, and wherein the preforms (2) are tempered with a radiant heater. 17. Method according to claim 13, wherein the step of selectively tempering the performs comprises contacting the performs with one of cooled and heated gas through at least one gas outlet opening (3). 18. Method according to claim 17, and the step of capturing the gas from the preforms (2) by the tempering device (1). 19. Method according to claim 17, and applying the cooled/heated gas by emitting the gas through a first gas outlet arrangement of a first tubular element (1a), surrounding the first tubular element (1a) with a second tubular element (1b) with a second gas outlet arrangement, whereby the gas outlet arrangements of the first (1a) and second tubular element (1b) partially overlap in a first relative position between the first and second tubular elements, and then moving the first tubular element (1a) and second tubular element (1b) into a second relative position in which the gas outlet arrangement of the first (1a) and second tubular element (1b) at least partially overlap to provide even tempering. 20. Method according to claim 19, and cooling the finished bottles by evenly tempering them. 21. Method according to claim 13, wherein the step of selectively tempering the performs comprises using a stretch rod (1) in the blow molding process (C). 22. Method according to claim 21, and the further steps of: inserting stretch rod (1) in the preheated preform (A), selectively cooling the preform (2), whereby the stretch rod (1) is not moved relative to the preform (2) (B), and stretching the stretch rod (1) in the preform (2) which is blow molded (C). 23. Device according to claim 5, wherein the cooled and heated gas is air. 24. Device according to claim 9, wherein the movement of the first (a) and second tubular element (2b) in relationship to each other is one of rotation and shifting. 25. Method according to claim 17 wherein the gas is air. |
3-phenoxy-4-pyridazinol derivatives and herbicide composition containing the same |
A compound represented by the formula: [wherein R1 represents a hydrogen atom, a halogen, atom, alkyl group, etc., R2 represents a hydrogen atom, a halogen atom, alkyl group, etc., R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a halogen atom, a substitutable alkyl group, a substitutable alkenyl group, alkynyl group, a substituteable cycloalkyl group, etc., or R3, R4, R5, R6 and R7 may form a ring which may be substituted, which is formed by the adjacent two of them with carbon atoms to which the respective substituents are bonded, m and n each independently represent 0 or 1.] a salt thereof, an ester derivative thereof and an agricultural chemical containing the same as an effective ingredient, and a herbicidal composition containing the compound and a second herbicidally active compound as effective ingredients. |
1. A compound represented by the formula: wherein R1 represents a hydrogen atom, a halogen atom, a C1 to C6 alkyl group, a C1 to C6 haloalkyl group, a C3 to C6 cycloalkyl group, a C2 to C6 alkenyl group, a cyano group, a C2 to C7 alkylcarbonyl group, a di(C1 to C6 alkyl)carbamoyl group, a phenyl group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a 5 or 6-membered heterocyclic group (the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may further contain 1 or 2 nitrogen atom(s)), a C1 to C6 alkoxy group, a phenoxy group which may be substituted (the substituent is a substituent selected from the following substituent Group A) or a 5- or 6-membered heterocycloxy group which may be substituted {the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may contain further contain 1 or 2 nitrogen atom(s), the substituent is a substituent(s) selected from the group consisting of a benzoyl group which may be substituted (the substituent is a substituent selected from the following substituent Group A) and a C1 to C6 alkyl group}, R2 represents a hydrogen atom, a halogen atom, a C1 to C6 alkyl group, a (C1 to C6 alkoxy)C1 to C6 alkyl group, a benzoyl group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a C2 to C7 alkoxycarbonyl group, a phenoxy group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a phenylthio group which may be substituted (the substituent is a substituent selected from the following substituent Group A) or a tri(C1 to C6 alkyl)silyl group, R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a halogen atom, a C1 to C6 alkyl group which may be substituted (the substituent is a substituent selected from the following substituent Group B), a C2 to C6 alkenyl group which may be substituted (the substituent is a cyano group or a nitro group), a C2 to C6 alkynyl group, a C3 to C6 cycloalkyl group which may be substituted (the substituent is a substituent selected from the following substituent Group C), a C4 to C10 bicycloalkyl group, a cyano group, a formyl group, a C2 to C7 alkylcarbonyl group, a benzoyl group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a carboxyl group, a C2 to C7 alkoxycarbonyl group, a carbamoyl group, a di(C1 to C6 alkyl)carbamoyl group, a phenyl group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a 3- to 6-membered heterocyclic group which may be substituted (the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may further contain 1 or 2 nitrogen atom(s), which may be fused with a benzene ring, the substituent is a substituent selected from the following substituent Group E), an amino group which may be substituted (the substituent is a substituent selected from the following substituent Group D), a nitro group, a hydroxyl group, a C1 to C6 alkoxy group, a C1 to C6 haloalkoxy group, a (C1 to C6 alkoxy) C1 to C6 alkoxy group, a phenoxy group which may be substituted (the substituent is a hydroxyl group or a pyridazinyloxy group substituted by a substituent(s) selected from the group consisting of a halogen atom and a C1 to C6 alkoxy group), a 5- to 6-membered heterocycloxy group which may be substituted (the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may contain further contain 1 or 2 nitrogen atom(s), the substituent is a substituent selected from the following substituent Group E), a phenylsulfonyloxy group which may be substituted (the substituent is a substituent selected from the following substituent Group A), a C1 to C6 alkylthio group, a C1 to C6 alkylsulfinyl group, a C1 to C6 alkylsulfonyl group or a tri(C1 to C6 alkyl)silyl group, or R3, R4, R5, R6 and R7 may form a 3- to 6-membered cyclic hydrocarbon group which may be substituted, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded (the cyclic hydrocarbon may be interrupted by the same or different 1 to 2 hetero atoms selected from the group consisting of a nitrogen atom, an oxygen atom and a sulfur atom, the substituent is a halogen atom, a C1 to C6 alkyl group, a hydroxy-C1 to C6 alkyl group, a C1 to C6 alkoxy group, an oxo group, a hydroxyimino group or a C1 to C6 alkoxyimino group, and when the C1 to C6 alkyl group is substituted, it may form another 3-membered ring by combining with the other C1 to C6 alkyl group or a carbon atom(s)in the cyclic hydrocarbon), m and n each independently represent 0 or 1, the substituent Group A is selected from the group consisting of a halogen atom, a C1 to C6 alkyl group, a C1 to C6 haloalkyl group, a C3 to C6 cycloalkyl group, a cyano group and a tri(C1 to C6 alkyl)silyl group, the substituent Group B is selected from the group consisting of a halogen atom, a C3 to C6 cycloalkyl group, a cyano group, a C2 to C7 alkylcarbonyl group, a C2 to C7 alkoxycarbonyl group, a phenyl group, a C1 to C6 alkoxy group, a C1 to C6 alkylthio group, a C1 to C6 alkylsulfinyl group, a C1 to C6 alkylsulfonyl group, a C1 to C4 alkylenedioxy group, a hydroxyimino group and a C1 to C6 alkoxyimino group, the substituent Group C is selected from the group consisting of a halogen atom, a C1 to C6 alkyl group which may be substituted (the substituent is a substituent selected from the substituent Group B), a C3 to C6 cycloalkyl group, a C2 to C6 alkenyl group, a cyano group, a C2 to C7 alkylcarbonyl group, a benzoyl group, a carboxyl group, a C2 to C7 alkoxycarbonyl group, a carbamoyl group, a di(C1 to C6 alkyl)carbamoyl group, a phenyl group which may be substituted (the substituent is a substituent selected from the above-mentioned substituent Group A), a 5 or 6-membered heterocyclic group (the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may contain further contain 1 or 2 nitrogen atom(s)), an amino group which may be substituted (the substituent is a substituent selected from the following substituent Group D), a nitro group, a hydroxyl group, a C1 to C6 alkoxy group, a C1 to C6 haloalkoxy group, a phenoxy group, a C1 to C6 alkylthio group, a phenylthio group, a C1 to C6 alkylsulfinyl group and a C1 to C6 alkylsulfonyl group, the substituent Group D is selected from the group consisting of a C1 to C6 alkyl group, a C2 to C7 alkylcarbonyl group, a C2 to C7 alkoxycarbonyl group, a di(C1 to C6 alkyl)carbamoyl group and a C1 to C6 alkylsulfonyl group, the substituent Group E is selected from the group consisting of a halogen atom, a C1 to C6 alkyl group, a C1 to C6 haloalkyl group, a hydroxyl group, a phenylsulfonyl group which may be substituted (the substituent is a substituent selected from the above-mentioned substituent Group A) and a di(C1 to C6 alkyl)sulfamoyl group], a salt thereof and an ester derivative thereof. 2. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group (the halogen atom is 1 to 3 fluorine atom(s)), a cyclopropyl group, a C2 to C3 alkenyl group, a cyano group, a C2 to C4 alkylcarbonyl group, a di(C1 to C3 alkyl)carbamoyl group, a phenyl group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a furyl group, a thienyl group, a C1 to C3 alkoxy group, a phenoxy group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 fluorine atom(s), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group} or a substituted pyrazolyloxy group (the substituent is a benzoyl group which is substituted by two chlorine atoms and two C1 to C3 alkyl groups). 3. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom, a bromine atom, a trifluoromethyl group or a cyano group. 4. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom or a bromine atom. 5. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom. 6. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R2 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C3 alkyl group, a (C1 to C3 alkoxy)C1 to C3 alkyl group, a benzoyl group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a C2 to C4 alkoxycarbonyl group, a phenoxy group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a phenylthio group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group} or a tri(C1 to C3 alkyl)silyl group. 7. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R2 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a methyl group, an ethoxycarbonyl group or a trimethylsilyl group. 8. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R2 is a hydrogen atom. 9. The compound, a salt thereof and an ester derivative thereof according to any of claims 1 to 8, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom, or a C3 to C4 cycloalkyl group, a C1 to C3 alkylthio group or a C1 to C3 alkoxyimino group), a C2 to C3 alkenyl group, a C2 to C3 alkynyl group, a C3 to C5 cycloalkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group, a cyano group, a C1 to C3 alkoxy group and a C1 to C3 alkylthio group), a C6 to C7 bicycloalkyl group, a cyano group, a C2 to C4 alkylcarbonyl group, a C2 to C4 alkoxycarbonyl group, a phenyl group which may be substituted {the substituent is a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group or a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a 5- to 6-membered heterocyclic group which may be substituted with 1 to 2 substituents which are the same or different {the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may further contain 1 or 2 nitrogen atom(s), the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group and a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a nitro group, a C1 to C3 alkoxy group, a C1 to C3 haloalkoxy group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a phenoxy group which may be substituted (the substituent is a pyridazinyloxy group which is substituted by a fluorine atom, a chlorine atom, a bromine atom or a C1 to C3 alkoxy group) or a C1 to C3 alkylthio group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2—, —CH2CH2CH2—, —CH(CH3)CH2CH2—, —CH2CH2CH2CH2—, —CH═CH—CH═CH—, —OCH2CH2—, —OCH═CH—, —OCH═C(CH3)—, —SCH═CH—, —N═CH—CH═CH—, —OCH2O—, —OCH2CH2O—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded. 10. The compound, a salt thereof and an ester derivative thereof according to any of claims 1 to 8, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted (the substituent is 1 to 3 fluorine atom(s), or a cyclopropyl group), a C3 to C4 cycloalkyl, group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is substituent selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C2 alkyl group, a cyclopropyl group and a C1 to C2 alkoxy group), a cyano group, a C2 to C3 alkoxycarbonyl group, a nitro group, a C1 to C3 alkoxy group or a trifluoromethoxy group, or R3, R4, R6 and R7 are a group represented by —CH2CH2CH2—, —CH(CH3)CH2CH2—, —OCH2CH2—, —OCH═CH— or which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. 11. The compound, a salt thereof and an ester derivative thereof according to any of claims 1 to 8, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a chlorine atom and C1 to C2 alkyl group), a cyano group or a C1 to C2 alkoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2— or —OCH═CH—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. 12. The compound, a salt thereof and an ester derivative thereof according to any of claims 1 to 8 or 13, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a methyl group, an ethyl group, an isopropyl group, a cyclopropyl group which may be substituted (the substituent is two chlorine atoms) or a methoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. 13. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein m and n are both 0. 14. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 fluorine atom(s), a cyclopropyl group, a C2 to C3 alkenyl group, a cyano group, a C2 to C4 alkylcarbonyl group, a di(C1 to C3 alkyl)carbamoyl group, a phenyl group which may be substituted {the substituent is the same or different 1 to 2 substituent(s) selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a furyl group, a thienyl group, a C1 to C3 alkoxy group, a phenoxy group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 fluorine atom(s), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group} or a substituted pyrazolyloxy group (the substituent is a benzoyl group which is substituted by two chlorine atoms and two C1 to C3 alkyl groups), R2 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C3 alkyl group, a (C1 to C3 alkoxy)C1 to C3 alkyl group, a benzoyl group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a C2 to C4 alkoxycarbonyl group, a phenoxy group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group}, a phenylthio group which may be substituted with 1 to 2 substituents which are the same or different {the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a cyclopropyl group, a cyano group and a tri(C1 to C3 alkyl)silyl group} or a tri(C1 to C3 alkyl)silyl group, R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom, or a C3 to C4 cycloalkyl group, a C1 to C3 alkylthio group or a C1 to C3 alkoxyimino group), a C2 to C3 alkenyl group, a C2 to C3 alkynyl group, a C3 to C5 cycloalkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group, a cyano group, a C1 to C3 alkoxy group and a C1 to C3 alkylthio group), a C6 to C7 bicycloalkyl group, a cyano group, a C2 to C4 alkylcarbonyl group, a C2 to C4 alkoxycarbonyl group, a phenyl group which may be substituted {the substituent is a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group or a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a 5- to 6-membered heterocyclic group which may be substituted with 1 to 2 substituents which are the same or different {the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may further contain 1 or 2 nitrogen atom(s), the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group and a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a nitro group, a C1 to C3 alkoxy group, a C1 to C3 haloalkoxy group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a phenoxy group which may be substituted (the substituent is a pyridazinyloxy group substituted by a fluorine atom, a chlorine atom, a bromine atom and C1 to C3 alkoxy group) or a C1 to C3 alkylthio group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2—, —CH2CH2CH2—, —CH(CH3)CH2CH2—, —CH2CH2CH2CH2—, —CH═CH—CH═CH—, —OCH2CH2—, —OCH═CH—, —OCH═C(CH3)—, —SCH═CH—, —N═CH—CH═CH—, —OCH2O—, —OCH2CH2O—, or which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, and m and n are both 0. 15. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom, a bromine atom, a trifluoromethyl group or a cyano group, R2 is a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a methyl group, an ethoxycarbonyl group or a trimethylsilyl group, R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted (the substituent is 1 to 3 fluorine atom(s), or a cyclopropyl group), a C3 to C4 cycloalkyl group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C2 alkyl group, a cyclopropyl group and a C1 to C2 alkoxy group), a cyano group, a C2 to C3 alkoxycarbonyl group, a nitro group, a C1 to C3 alkoxy group or a trifluoromethoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2—, —CH(CH3)CH2CH2—, —OCH2CH2—, —OCH═CH— or which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom, and m and n are both 0. 16. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom or a bromine atom, R2 is a hydrogen atom, R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a chlorine atom and C1 to C2 alkyl group), a cyano group or a C1 to C2 alkoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2— or —OCH═CH—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom, and m and n are both 0. 17. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein R1 is a chlorine atom, R2 is a hydrogen atom, R3, R4, R5 R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a methyl group, an ethyl group, an isopropyl group, a cyclopropyl group which may be substituted (the substituents are two chlorine or a methoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2—, which is formed by two adjacent members of them R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom, and m and n are both 0. 18. The compound, a salt thereof and an ester derivative thereof according to claim 1, wherein the compound is selected from the group consisting of 6-chloro-3-(2-iodophenoxy)-4-pyridazinol, 6-chloro-3-(2-methylphenoxy)-4-pyridazinol, 6-chloro-3-(2-cyclopropylphenoxy)-4-pyridazinol, 6-chloro-3-(2,3-dihydro-1H-inden-4-yloxy)-4-pyridazinol, 3-(1-benzofuran-7-yloxy)-6-chloro-4-pyridazinol, 6-chloro-3-(2-methoxy-5-methylphenoxy)-4-pyridazinol, 6-chloro-3-(2-chloro-6-cyclopropylphenoxy)-4-pyridazinol, 3-(2-bromo-6-methylphenoxy)-6-chloro-4-pyridazinol, 6-chloro-3-(2-cyclopropyl-6-methylphenoxy)-4-pyridazinol and 6-chloro-3-(2-cyclopropyl-3,5-dimethylphenoxy)-4-pyridazinol. 19. An agricultural chemical composition which comprises the compound, a salt thereof and an ester derivative thereof according to claim 1 as an effective ingredient in combination with a carrier. 20. A herbicidal composition which comprises (i) at least one 3-phenoxy-4-pyridazinol compound selected from the group consisting of the compound, a salt thereof and an ester derivative thereof according to claim 1, and (ii) at least one herbicidally active compound selected from the group consisting of 4-(2,4-dichlorobenzoyl)-1,3-dimethyl-5-pyrazolyl-p-toluenesulfonate, 2-[4-(2,4-dichlorobenzoyl)-1,3-dimethylpyrazol-5-yloxy]acetophenone, 2-[4-(2,4-dichloro-m-toluoyl)-1,3-dimethylpyrazol-5-yloxy]-4′-methylacetophenone, 5-cyclopropyl-1,2-oxazol-4-yl α-α-α-trifluoro-2-mesyl-p-tolyl ketone, 2-(2-chloro-4-mesylbenzoyl)cyclohexan-1,3-dione, 2-(4-mesyl-2-nitrobenzoyl)cyclohexan-1,3-dione and 4-chloro-2-(methylsulfonyl)phenyl 5-cyclopropyl-4-isoxazolyl ketone, as effective ingredients. 21. The herbicidal composition according to claim 20, wherein the herbicidally active compound is 4-(2,4-dichlorobenzoyl)-1,3-dimethyl-5-pyrazolyl-p-toluenesulfonate. 22. 2-Cyclopropyl-6 methylphenol. 23. The compound, a salt thereof and an ester derivative thereof according to claim 13, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom, or a C3 to C4 cycloalkyl group, a C1 to C3 alkylthio group or a C1 to C3 alkoxyimino group), a C2 to C3 alkenyl group, a C2 to C3 alkynyl group, a C3 to C5 cycloalkyl group which may be substituted with 1 to 3 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group, a cyano group, a C1 to C3 alkoxy group and a C1 to C3 alkylthio group), a C6 to C7 bicycloalkyl group, a cyano group, a C2 to C4 alkylcarbonyl group, a C2 to C4 alkoxycarbonyl group, a phenyl group which may be substituted {the substituent is a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group or a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a 5- to 6-membered heterocyclic group which may be substituted with 1 to 2 substituents which are the same or different {the heterocycle contains one nitrogen atom, oxygen atom or sulfur atom in the ring, and may further contain 1 or 2 nitrogen atom(s), the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C3 alkyl group and a C1 to C3 haloalkyl group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom)}, a nitro group, a C1 to C3 alkoxy group, a C1 to C3 haloalkoxy group having 1 to 3 halogen atoms which are the same or different (the halogen atom is selected from the group consisting of a fluorine atom, a chlorine atom and a bromine atom), a phenoxy group which may be substituted (the substituent is a pyridazinyloxy group which is substituted by a fluorine atom, a chlorine atom, a bromine atom and a C1 to C3 alkoxy group) or a C1 to C3 alkylthio group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2—, —CH2CH2CH2—, —CH(CH3)CH2CH2—, —CH2CH2CH2CH2—, —CH═CH—CH═CH—, —OCH2CH2—, —OCH═CH—, —OCH═C(CH3)—, —SCH═CH—, —N═CH—CH═CH—, —OCH2O—, —OCH2CH2O—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded. 24. The compound, a salt thereof and an ester derivative thereof according to claim 13, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C4 alkyl group which may be substituted (the substituent is 1 to 3 fluorine atom(s), or a cyclopropyl group), a C3 to C4 cycloalkyl group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a fluorine atom, a chlorine atom, a bromine atom, a C1 to C2 alkyl group, a cyclopropyl group and a C1 to C2 alkoxy group), a cyano group, a C2 to C3 alkoxycarbonyl group, a nitro group, a C1 to C3 alkoxy group or a trifluoromethoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2—, —CH (CH3) CH2CH2—, —OCH2CH2—, —OCH═CH— or which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. 25. The compound, a salt thereof and an ester derivative thereof according to claim 13, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a C1 to C3 alkyl group, a C3 to C4 cycloalkyl group which may be substituted with 1 to 2 substituents which are the same or different (the substituent is selected from the group consisting of a chlorine atom and C1 to C2 alkyl group), a cyano group or a C1 to C2 alkoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2— or —OCH═CH—, which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. 26. The compound, a salt thereof and an ester derivative thereof according to claim 13, wherein R3, R4, R5, R6 and R7 each independently represent a hydrogen atom, a fluorine atom, a chlorine atom, a bromine atom, an iodine atom, a methyl group, an ethyl group, an isopropyl group, a cyclopropyl group which may be substituted (the substituent is two chlorine atoms) or a methoxy group, or R3, R4, R5, R6 and R7 are a group represented by —CH2CH2CH2— which is formed by two adjacent members of R3, R4, R5, R6 and R7 with carbon atoms to which respective substituents are bonded, provided that R3 is not a hydrogen atom. |
<SOH> BACKGROUND ART <EOH>In Chemical Pharmaceutical Bulletin, 1972, vol. 20, No. 10, pp. 2191-2203, 3-(2-allylphenoxy)-6-chloro-4-methoxypyridazine has been disclosed but a 3-phenoxy-4-pyridazinol compound having a hydroxyl group at the 4-position of the pyridazine has not been disclosed, and there is no description about a herbicide. In Journal of the Chemical Society: Perkin Transaction I, 1975, No. 6, pp. 534-538, 3-(2-hydroxyphenoxy)-4-methoxypyridazine and 6-chloro-3-(2-hydroxyphenoxy)-4-methoxypyridazine has been disclosed but a 3-phenoxy-4-pyridazinol compound having a hydroxyl group at the 4-position of the pyridazine has not been disclosed, and there is no description about a herbicide. In U.S. Pat. No. 5,559,080, a 3-(phenoxy which may be substituted)pyridazine compound having a haloalkylphenoxy group at the 4-position of the pyridazine has been disclosed but a 3-phenoxy-4-pyridazinol compound having a hydroxyl group at the 4-position of the pyridazine has not been disclosed. Also, in the 3-(phenoxy which may be substituted)pyridazine compound having a haloalkylphenoxy group at the 4-position of the pyridazine, an oxygen atom bonded to the 4-position of the pyridazine is bonded by a benzene ring, and its herbicidal activity was insufficient. Also, at present, a number of herbicides have been practically used as a herbicide for a paddy field, and widely been used for general purpose as a single agent and a mixed agent. However, there are many kinds of paddy field weeds, and germination and growth period of the respective weeds are not uniform, in particular, occurrence of perennial weeds ranges for a long period of time. Thus, it is extremely difficult to prevent from and kill all weeds with one time spread of a herbicide. Accordingly, as a herbicide, an appearance of a chemical which can kill many kinds of weeds including annual weeds and perennial weeds, that is, which has a wide weed-killing spectrum, is effective for already grown weeds, preventing and killing effects of weeds of which can be maintained for a certain period of time, and has high safety to paddy rice has earnestly been desired. Also, as upland herbicides, a number of herbicides have now been commercially available and practically used, but there are many kinds of weeds to be prevented, and occurrence thereof ranges for a long period of time, so that a herbicide which has higher herbicidal effects, has broad weed-killing spectrum, and causes no chemical damage to crops has been desired. One of the effective ingredient of the herbicidal composition of the present invention (hereinafter referred to as a second herbicidally active compound), 4-(2,4-dichlorobenzoyl)-1,3-dimethyl-5-pyrazolyl-p-toluenesulfonate [hereinafter referred to as Compound A. General name: Pyrazolate], 2-[4-(2,4-dichlorobenzoyl)-1,3-dimethylpyrazol-5-yloxy]acetophenone [hereinafter referred to as Compound B. General name: Pyrazoxyfen], 2-[4-(2,4-dichloro-m-toluoyl)-1,3-dimethylpyrazol-5-yloxy]-4′-methylacetophenone (hereinafter referred to as Compound C. General name: Benzofenap], 5-cyclopropyl-1,2-oxazol-4-yl α,α,α-trifluoro-2-mesyl-p-tolyl ketone [hereinafter referred to as Compound D. General name: Isoxaflutole], 2-(2-chloro-4-mesylbenzoyl)cyclohexan-1,3-dione [hereinafter referred to as Compound E. General name: sulcotrione], 2-(4-mesyl-2-nitrobenzoyl)cyclohexan-1,3-dione [hereinafter referred to as Compound F. General name: mesotrion] and 4-chloro-2-(methylsulfonyl)phenyl 5-cyclopropyl-4-isoxazolyl ketone [hereinafter referred to as Compound G. General name: Isoxachlortole] are each conventionally known herbicidal compound, and each described in The Pesticide Manual 11th Edition, pp. 1049 to 1050, Ibid. pp. 1054 to 1055, Ibid. pp. 111 to 112, The Pesticide Manual, 12th Edition p. 563, Ibid. p. 848, Ibid. p. 602 and EP 470 856(1990). These compounds have high effects against annual broad-leaved weeds and a part of perennial weeds, but their effects against rice plant weeds or a part of perennial weeds are not necessarily sufficient. |
Secure gateway with proxy service capability servers for service level agreement checking |
The present invention describes a gateway in an Open Service Access (OSA) network where Service Level Agreement (LSA) checks are performed by a Framework (15) on a Central Gateway node (1). A distinction is made between applications (10) that can be trusted, like applications provided by the same firm as the gateway, and other applications (12) that are not trusted for security reasons. Access request coming front the applications for accessing Service Capability Servers (SCSs) (4, 5) are checked by the framework (15). Now the trusted applications (10) can get direct access to the Service Capability Servers (4, 5), but the untrusted applications (12) are only allowed to access so-called proxy SCSs (9) on the Central Gateway node (1). The proxy SCSs (9) have the same interface as the SCSs (4, 5) running on the distant nodes (2, 3), and are downloaded by the Framework (15) from the distant SCS nodes (2, 3) to the Central Gateway node (1) during an initialization phase. |
1. A gateway node for an open service access network between one or more applications and one or more external service capability servers on service capability server nodes, comprising a a database for storing service legal agreements associated with one or more of said applications; a proxy service capability server for communicating with one or more of said applications and said one or more external service capability servers; means for perform security checks on requests from applications to get access to one or more of the external service capability servers using service level agreements stored in said database; classify the requesting applications as trusted and untrusted applications, and give a trusted application direct access to said external service capability servers but give an untrusted application only access to proxy service capability server running on the gateway node and said untrusted application communicating with said one or more external service capability servers through said proxy service capability server. 2. A gateway node according to claim 1, wherein said means for performing security checks is arranged to initiate downloading of application program interface functionality from said service capability servers to said gateway node forming said proxy service capability servers. 3. Method of a gateway node comprising the steps of: (a) receiving requests from an application to access one or more service capability servers external to said gateway node (b) performing security checks on said requests using service level agreements, stored in a database; (c) downloading of application program interface functionality from said service capability servers to the gateway node forming proxy service capability servers; (d) classifying the application into either a trusted or an untrusted application; (e) requesting said external service capability server to create an object instance implementing said application program interface functionality; (f) sending a reference to said object instance to said application if it is a trusted application; (g) sending a reference to said proxy service capability servers to said application if it is an untrusted application; (h) operating said proxy service capability servers to enable controlled communications between said untrusted application and said service capability server. 4. Method of a gateway node, according to claim 3, wherein said proxy service capability servers are formed on said gateway node every time said one or more applications need access to said service capability servers. 5. Method of a gateway node, according to claim 3, wherein said proxy service capability servers are formed on said gateway node during an installation time of said gateway node. 6. A gateway node for providing communication between one or more applications and one or more service capability servers, comprising: (a) means for receiving requests from an application to access one or more service capability servers external to said gateway node; (b) means for performing security checks on said requests using service level agreements, stored in a database; (c) means for downloading of application program interface functionality from said service capability servers to the gateway node forming proxy service capability servers; (d) means for classifying the application into either a trusted or an untrusted application; (e) means for requesting said external service capability server to create an object instance implementing said application program interface functionality; (f) means for sending a reference to said object instance to said application if it is a trusted application; (g) means for sending a reference to said proxy service capability servers to said application if it is an untrusted application; (h) means for operating said proxy service capability servers to enable controlled communications between said untrusted application and said service capability server. 7. The gateway node according to claim 6, wherein said proxy service capability servers are formed on said gateway node every time said one or more applications need access to said service capability servers. 8. The gateway node according to claim 6, wherein said proxy service capability servers are formed on said gateway node during an installation time of said gateway node. |
<SOH> FIELD OF THE INVENTION <EOH>The present invention relates to Open Service Access (OSA) for the implementation of a third generation wireless phone system called Universal Mobile Telecommunications System (UMTS). The invention particularly relates to securing access to Service Capability Servers (SCSs). |
<SOH> SUMMARY OF THE INVENTION <EOH>The problem to be solved by the present invention is to provide a system and method that overcome the disadvantages of the prior art approaches. The present invention relates to a gateway node for implementing a gateway in an open service access network between one or more applications and one or more external service capability servers on service capability server nodes, comprising a framework, characterised in that said framework performs security checks on requests from applications to get access to nore or more of the external service capability servers using service level agreements, stored in a database. In the present invention the physical and logical gateway approach are combined to benefit from both prior art approaches. Futhermore, the present invention relates to a gateway nodes as described above, wherein said framework classifies the applications in trusted and untrusted applications. Moreover, the present invention relates to a gateway node as described above, wherein said Framework allows a trusted application direct access to said service capability servers but allows a untrusted application only access to proxy service capability servers running on the gateway node. Also, the present invention relates to a gateway node as described above, wherein said framework initiates downloading of application program interface functionality from said service capability servers to said gateway node forming said proxy service capability servers. Futhermore, the present invention relates to a method for implementing a gateway node as described above, comprising the steps of: (a) receiving a request from an application to access a service capability server external to said gateway node, characterised in that the method also comprises the steps of: (b) downloading of application program interface functionality from said service capability servers to the gateway node forming a proxy service capability servers; (c) classifying the application into either a trusted or an untrusted application; (d) requesting said external service capability server to create an object instance implementing required application program interface functionality; (e) sending a reference to said object instance to said application if it is a trusted application; (f) sending a reference to said proxy SCS to said application if it is an untrusted application; (g) operating said proxy service capability servers to enable controlled communications between said untrusted application and said service capability server. In the present invention on the fly software can be provided to a node in the network that can deal with the checking of the Service Level Agreement. This keeps the actual SCS from doing these checks ad therefore trusted applications are not bothered with the SLA checking overhead. Meanwhile untrusted applications do not have direct access to the actual SCSs and have to go through one specific node, which is beneficial from a security point of view. |
System and method for starting a catalytic reactor |
A system and method is provided for starting a catalytic reactor supplied with an oxygen-containing reactant gas, such as air, and a vaporized liquid fuel comprising carbon and hydrogen, such as methanol. The temperature difference between the temperature in the inlet area of the catalytic reactor and the temperature in the outlet area of the catalytic reactor is monitored, and the supply of the liquid fuel is adjusted based on the temperature difference, while the reactant gas is supplied to the catalytic reactor continuously. |
1. A method of starting a catalytic reactor, comprising: supplying the reactor with a reactant gas stream comprising oxygen and an atomized liquid fuel, determining a temperature difference between a reactor inlet temperature and a reactor outlet temperature, and adjusting the supply of the liquid fuel based on the temperature difference. 2. The method of claim 1, wherein the reactant gas is air. 3. The method of claim 1, wherein the liquid fuel comprises a hydrocarbon. 4. The method of claim 1, wherein the supply of liquid fuel is stopped when the reactor outlet temperature is less than the reactor inlet temperature. 5. The method of claim 4, further comprising restarting the supply of the liquid fuel once the reactor outlet temperature is greater than the reactor inlet temperature. 6. The method of claim 4, further comprising restarting the supply of the liquid fuel after a predetermined period. 7. The method of claim 1, further comprising introducing thermal energy into the reactor, and wherein the reactor outlet temperature used in determining the temperature difference is adjusted to account for the temperature increase resulting from the introduction of the thermal energy. 8. The method of claim 1, further comprising introducing thermal energy into the gas upstream of the reactor, and wherein the determined temperature difference takes into account the temperature increase resulting from the introduction of thermal energy. 9. The method of claim 1, wherein the reactor is a component of a fuel cell system. 10. The method of claim 9, wherein the fuel cell system is a component of a motor vehicle. 11. A system for starting a catalytic reactor comprising: a gas supply passage configured to supply a gas stream to an inlet port of the reactor, a liquid fuel supply passage comprising an atomizer to introduce an atomized liquid fuel into the gas supply passage; an outlet passage for directing fluid from an outlet port of the reactor, at least one temperature sensor disposed adjacent to each of the inlet and the outlet ports, and a control valve disposed in the liquid fuel supply passage and couple to receive an input signal from the temperature sensors. 12. The system of claim 11, further comprising a heater. 13. The system of claim 12, wherein the heater is disposed in the gas supply passage. 14. The system of claim 12, wherein the heater is disposed in the reactor. 15. The system of claim 11, wherein the gas supply passage comprises a turn of approximately 90° upstream of the inlet port, and an accumulated liquid fuel removal passage disposed adjacent to the turn. 16. The system of claim 11, wherein the system is a component of a fuel cell system for a motor vehicle. |
<SOH> BACKGROUND OF THE INVENTION <EOH>1. Field of the Invention The invention concerns a system and method for starting a catalytic reactor. 2. Description of the Related Art During the low-temperature start-up of catalytic reactors, such as those used in a motor vehicle fuel cell system with a gas generation system, the reactants must be available in gaseous form. Often, at least one of the reactants is a liquid under ambient conditions. The liquid reactant is atomized, such as by using a nozzle or a similar device, into a gaseous second reactant stream flowing into the catalytic reactor. In certain circumstances, for example at start-up, the liquid reactant is typically not evaporated entirely In the gaseous second reactant stream, since during startup of the reactor the stream is typically cold. Since the temperature of the catalytic reactor itself is far below the operating temperature, the evaporation can also not be completed in the catalytic reactor, before the reaction between the vaporized liquid reactant and the gaseous reactant takes place. A severely disadvantageous result of this is that at least part of the reactant stream mixture enters the reactor in liquid form. If the reactor employs a porous catalyst support, then liquid reactant can accumulate in this catalyst support prior to its conversion in the reactor. When the catalytic reaction commences, then, as a result of accumulation of liquid reactant, the catalytic reaction takes place in the reactor at a far greater concentration of the liquid reactant than that intended (i.e. greater than the desired concentration of liquid reactant which was introduced into the gaseous reactant stream). This may be detrimental to the catalyst, since it can overheat in some areas due to the very high concentration of reactant. Moreover, the presence of liquid reactant in the catalytic reactor can slow reactor start-up, as the presence of the liquid reactant in the catalyst support can block access of second reactant to the catalyst. This significantly impedes the start of the desired reaction. Accordingly, there is a need for a system and method for starting a catalytic reactor operating on a vaporized liquid reactant, and for heating the reactor to the desired operating temperature (particularly from a low temperature), in as short a time as possible, whereby the emissions of unreacted reactants and by-products, and the degradation of the catalytic material are reduced. |
<SOH> SUMMARY OF THE INVENTION <EOH>Using the present system and method, a catalytic reactor is started by supplying a liquid fuel, such as methanol, into a continuously circulated gaseous reactant stream, such as an air stream (which supplies the necessary oxygen), while monitoring the temperature in the area of both the reactor inlet and outlet. The liquid fuel is atomized in a feed line upstream of the catalytic reactor and is partially evaporated in the gaseous reactant stream. This results in cooling of the gaseous mixture, which can be detected by a drop in the reactor inlet temperature. When the reactor is operational, the gaseous portion of the fuel reacts with the gaseous reactant, on the catalytically active surface of the catalytic reactor, which may for example have been applied as a coating onto a porous catalyst support. Additional amounts of the liquid fuel can be evaporated using the thermal energy that is generated by the catalytic reaction and can subsequently be converted in the catalytic reactor. If there is a reduction in the activity of the catalyst, or if there is an insufficient concentration of fuel, the gaseous mixture flows through the catalytic reactor without any significant amount of conversion taking place. If liquid fuel accumulates in the porous catalyst support due to capillary action, preventing the gaseous reactants from contacting the catalyst, the desired reaction will be inhibited and the catalyst support will cool to the temperature of the incoming mixture. This temperature drop will result in a drop in the reactor outlet temperature. Despite the continued supply of liquid fuel into the gaseous reactant stream, the catalytic reactor will not get started, since the temperature will continue to drop and the catalytic material becomes flooded with the liquid fuel. As this occurs, the reactor outlet temperature approaches the reactor inlet temperature. In the present system and method, the temperature difference between the reactor inlet and the reactor outlet is monitored and, if the temperature in the reactor fails to increase, the supply of liquid fuel is reduced or shut off, while the gaseous reactant stream continues to be supplied to the reactor. Subsequently, the catalyst support of the catalytic reactor will be heated slightly by the entering gaseous reactant stream, into which no, or a much smaller amount of, liquid fuel is being introduced. This allows the liquid fuel accumulated in the catalyst support to be at least partially evaporated by the thermal energy in the incoming gaseous reactant stream. Once the fuel is present in the reactor in gaseous form, it reacts with the reactive component of the gaseous reactant stream. As soon as the reaction commences in local areas of the catalyst support, the heat that is produced by that (exothermic) reaction spreads out, evaporating liquid fuel which has accumulated in the surrounding catalyst support. Eventually, the reaction propagates throughout the entire catalytic reactor. The start of the reaction can be defected by means of an increase in the reactor outlet temperature. As soon as a specific reactor outlet temperature has been reached or the difference between the reactor outlet temperature and reactor inlet temperature has become sufficiently positive, the supply of liquid fuel can then be commenced or increased. The evaporation of the accumulated liquid fuel into the gas stream should lead to a cooling of the catalytic reactor. If this cooling does not take place, i.e. if the temperature difference between the reactor inlet temperature and the reactor outlet temperature rises to zero or a value greater than zero, this is an indication that there is no remaining liquid fuel in the catalyst support that could evaporate. This state can also be used to trigger starting or increasing the supply of liquid fuel into the gaseous reactant stream, so that the described sequence can start from the beginning. The entire process can be repeated as often as is necessary to achieve a successful start-up of the catalytic reactor. The advantage of the present system and method is that it can be implemented very easily. The temperature difference is dependent on the amount of liquid fuel supplied and evaporated (because of the cooling which occurs as a result of the evaporation of the fuel into the gaseous reactant stream) and on the reaction of the mixture in the catalytic reactor, which generates heat. The supply of liquid fuel into the gaseous reactant stream is adjusted based on the monitored temperature difference. The apparatus required is very simple, as only one additional temperature sensor is required. The continuous supply and circulation of the gaseous reactant stream through the reactor, which leads to unreacted liquid fuel being discharged from the catalytic reactor, can reduce catalyst degradation since there will not be excessively high local concentrations of the fuel and less tendency for local overheating to occur. If the reactants used are air and a liquid fuel that contains carbon and hydrogen, then this reduction in locally excessive concentrations of liquid fuel will prevent, or at least reduce, hydrocarbon and carbon monoxide emissions which would be the result of a local combustion with a lambda value of λ<1, i.e. fuel excess. As a further advantage, the present system and method offers significant time saving in the start-up of such a catalytic reactor when compared to sequence that includes aborting the start-up procedure, followed by a complete purging of the system, and a re-start. These and other aspects will be evident upon reference to the attached Figures and following detailed description. |
Novel device |
A device for delivering an oral healthcare substance to the teeth, gingival and/or mucosal tissues comprising a strip of an orally acceptable flexible material with an oral healthcare substance deposited onto it or impregnated into its bulk, capable of adhesion to a tooth surface but with the adhesion function being provided independent of the oral healthcare substance. A preferred device comprises a strip of a plastically deformable material, to which is attached a layer of an absorbent material, with a peroxide-containing tooth whitening gel on the layer of absorbent material. |
1. A delivery device for delivering an oral healthcare substance to the oral surfaces of the teeth, gingival and/or mucosal tissues, comprising; a strip of an orally acceptable flexible material, having a strip surface capable of being applied to a tooth surface and/or adjoining soft tissue, the strip having an oral healthcare substance deposited upon a strip surface thereof, and/or impregnated into its bulk, the substance being capable of transference from the strip surface to the tooth surface and/or adjoining soft tissue, the strip being capable of adhesion to a tooth surface and/or adjoining soft tissue to which the strip is applied, the adhesion function being provided independent of the oral healthcare substance. 2. A delivery device according to claim 1 wherein the strip comprises a plastically deformable material. 3. A device according to claim 2 wherein the plastically deformable material comprises a wax. 4. A device according to claim 2 wherein the plastically deformable material when in the form of the strip may be deformed by the user using finger or hand pressure to fit the device to the overall shape of the user's teeth. 5. A device according to claim 4 made of a plastically deformable material such that physical conformation of the plastically deformable material to the shape of the teeth causes attachment of the device to the teeth. 6. A device according to claim 4 wherein the material is capable of being plastically deformed so that the device can be folded or bent over the user's teeth from the front to the back surface of the teeth, so as to be in contact with both the front and back surfaces. 7. A delivery device according claim 1, wherein the strip comprises an inner carrier layer able to carry the substance and to be placed in contact with a user's teeth, and an outer barrier layer. 8. A delivery device according to claim 7 which comprises a barrier layer of a plastically deformable material, to at least one surface of which is attached a carrier layer of an absorbent material, an oral healthcare substance being on the carrier layer of absorbent material. 9. A device according to claim 8 wherein the absorbent material is an absorbent fabric. 10. A device according to claim 9 wherein the fabric is non-woven. 11. A device according to claim 10 wherein the non-woven fabric is a polypropylene-viscose blend. 12. A device according to claim 1, wherein the oral care substance and an adhesive material are deposited in separate discrete locations in relation to the strip surface. 13. A device according to claim 12 wherein the adhesive is deposited in places on the strip surface that enable part of the strip to stick to an oral surface adjacent to a tooth surface, so that another part of the strip on which the substance is deposited or into which it is impregnated contacts the tooth surface. 14. A device according to claim 12 wherein the oral healthcare substance and adhesive are deposited on the surface in respective discrete spots or patches or lines on the surface. 15. A device according to claim 14 wherein the oral healthcare substance is deposited in one or more patch bordered partly or completely surrounded by a border of the adhesive. 16. A device according to claim 11 wherein the adhesive and oral healthcare substance are deposited in separate discrete layers on the surface of the strip. 17. A device according to claim 1, wherein the strip material is inherently adherent to a tooth surface. 18. A device according to claim 1, wherein mechanical adhesive means provides an adhesive function, either alone or in combination with any other adhesive means. 19. A device according to claim 1, wherein the oral care substance and/or adhesive are encapsulated. 20. A device according to claim 1, wherein the oral care substance and/or adhesive are provided in granules which may release their content under the influence of the mouth environment. 21. A device according to claim 19 wherein said granules or capsules are covered by a porous membrane layer. 22. A device according to claim 1, comprising an outer backing layer and an inner contact layer to be placed adjacent a tooth or other oral tissue surface, the inner layer being porous and the oral healthcare substance being provided between the inner and outer layers. 23. A delivery device according to claim 1, marked with one or more visible symbol. 24. A delivery device according to claim 23 wherein a cover layer is applied over the one or more symbol. 25. A device according to claim 23 which comprises a barrier layer of a plastically deformable material, to at least one surface of which is attached a carrier layer of an absorbent material, an oral healthcare substance on the carrier layer of absorbent material, and marked with one or more visible symbol applied to the surface of the plastically deformable material opposite to the surface on which is attached the layer of an absorbent material, and having a cover layer over the one or more symbol. 26. A device according to claim 25 wherein the cover layer is the same material as the plastically deformable material so that the symbol is in effect embedded in the plastically deformable material. 27. A delivery device according to claim 1, wherein the oral care substance is a tooth whitening substance. 28. A delivery device according to claim 1, wherein the oral care substance is in the form of a gel. 29. A delivery device according to claim 27 wherein the oral healthcare substance is a peroxide-containing gel. 30. A device according to claim 1, being of elongate shape of a length sufficient that when placed against the front surface of the teeth of a user it extends across a plurality of teeth, and of sufficient width that when placed against the user's teeth it extends from the gumline at least to the crowns of the front teeth distant from the gumline. 31. A device according to claim 30 wherein the width is such that in an unfolded state the strip has an unfolded width greater than the height of the teeth from the gumline to the crown, and at least part of the strip may be folded about a substantially longitudinal fold axis so as to bend or fold over the crowns and contact the crowns and rear surfaces of the user's teeth. 32. A device according to claim 30 being substantially rectangular with concavely curved long sides. 33. A device according to claim 30 substantially rectangular with convexly bowed long sides, or of a generally rectangular shape but with a concave curved long side or a concave indentation in a long side. 34. A device according to claim 1 having a tab extending from a long side. 35. A device according to claim 34 wherein at least part of the strip may be folded into a folded cross section substantially of a “U” or “V” shape, with two limbs linked at a fold axis each limb with an inward surface facing into the “bite” of the “U” or “V”, with the oral healthcare substance present on this inward, tooth-contacting surface. 36. A method of treating teeth with a device according to claim 1 in which the device is applied manually to a user's tooth surface(s), with the length dimension of the strip aligned with the line of the user's teeth, the substance-bearing surface in contact with the front surface of the teeth, and the device is pressed against the front surface of the teeth, leaving the device in contact with the user's teeth for a sufficient length of time for the oral healthcare substance to have its effect, then removing the device. 37. A process for making a device according to claim 8 comprising bringing the absorbent material into contact with the surface of the strip and applying pressure, then applying the oral healthcare substance to the absorbent material. 38. A process according to claim 37 comprising; providing a dental wax in the form of a sheet, optionally cleaning the surface of the sheet of the wax material, printing a visible symbol upon a first surface of a first sheet of the wax material, laminating a second, cover sheet of the wax material to the first surface of the first sheet, to sandwich the printed symbol between the first and second sheets, sizing the laminated sheets by compressing them to a suitable thickness, attaching the layer of absorbent fabric to the laminated wax strip, cutting the sheet of laminated material to shape, applying the oral healthcare substance to the layer of absorbent material. 39. A device according to claim 3-wherein the plastically deformable material when in the form of the strip may be deformed by the user using finger or hand pressure to fit the device to the overall shape of the user's teeth. 40. A device according to claim 5 wherein the material is capable of being plastically deformed so that the device can be folded or bent over the user's teeth from the front to the back surface of the teeth, so as to be in contact with both the front and back surfaces. 41. A delivery device according to claim 6 wherein the strip comprises an inner carrier layer able to carry the substance and to be placed in contact with a user's teeth, and an outer barrier layer. 42. A device according to claim 11 wherein the oral care substance and an adhesive material are deposited in separate discrete locations in relation to the strip surface. 43. A device according to claim 13 wherein the oral healthcare substance and adhesive are deposited on the surface in respective discrete spots or patches or lines on the surface. 44. A device according to claim 20 wherein said granules or capsules are covered by a porous membrane layer. 45. A device according to claim 24 which comprises a barrier layer of a plastically deformable material, to at least one surface of which is attached a carrier layer of an absorbent material, an oral healthcare substance on the carrier layer of absorbent material, and marked with one or more visible symbol applied to the surface of the plastically deformable material opposite to the surface on which is attached the layer of an absorbent material, and having a cover layer over the one or more symbol. 46. A delivery device according to claim 28 wherein the oral healthcare substance is a peroxide-containing gel. 47. A device according to claim 31 being substantially rectangular with concavely curved long sides. 48. A device according to claim 1 substantially rectangular with convexly bowed long sides, or of a generally rectangular shape but with a concave curved long side or a concave indentation in a long side. 49. A device according to claim 33 having a tab extending from a long side. |
Glycoconjugates of sialic acid derivates, methods for their production and use thereof |
The present invention relates to glycoconjugates containing a sialic acid derivate of general formula (I) and wherein the sialic acid derivate of general formula (I) is conjugated to a mono-, di- or oligosaccharide with up to 40 glycosidically linked, optionally branched sugar residues representing furanose and/or pyranose rings, which are linked N- or O-glycosidically to the polypeptide. The sialic derivatives of general formula (I) are useful for producing pharmaceutical compositions for immunosuppression, cell protection, stimulation of hematopoesis regulation of hormonal secretion and hormonal activation. |
1. Glycoconjugates obtained by conjugating a sialic acid derivate of general formula I to a mono-, di- or oligosaccharide with up to 40 glycosidically linked, optionally branched sugar residues representing furanose and/or pyranose rings, which are linked N- or O-glycosidically to a polypeptide wherein R1 represents hydrogen or lower alkyl up to 5 carbon atoms, which may be branched, unbranched, acyclic, alicyclic or cyclic, R2 is acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic, R3 is a halogen atom, a methylsulfide group, a methylsulfate group or acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic, R4 represents, independently from each other, hydrogen, acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic, R5 is hydrogen or a halogen atom R6 represents hydrogen, acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic and wherein the sialic acid derivate of general formula I is conjugated to said mono-, di- or oligosaccharide via R6. 2. Glycoconjugates according to claim 1, wherein R1 represents hydrogen or lower alkyl up to 3 carbon atoms, which may be branched, unbranched or cyclic, R2 is acetyl, R3 represents, independently from each other, hydrogen or acetyl, R4 represents, independently from each other, hydrogen or acetyl and R5 is hydrogen. 3. Glycoconjugates according to claim 1, wherein R1 represents hydrogen, methyl or ethyl, R2 is acetyl, R3 represents, independently from each other, hydrogen or acetyl, R4 represents, independently from each other, hydrogen or acetyl and R5 is hydrogen. 4. Glycoconjugates according to claim 1, wherein the sialic acid derivate to be incorporated is selected from 5-N-acetyl-9-deoxy-neuraminic acid, 5-N-acetyl-9-deoxy-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-amino-9-deoxy-neuraminic acid, 5-N-acetyl-9-amino-9-deoxy-neuraminic acid peracetylate, 5-N-acetyl-9-amino-9-deoxy-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-amino-9-deoxy-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-acetamido-9-deoxy-neuraminic acid, 5-N-acetyl-9-acetamido-9-deoxy-neuraminic acid peracetylate, 5-N-acetyl-9-acetamido-9-deoxy-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-acetamido-9-deoxy-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-aminoacetamido-9-deoxy-neuraminic acid, 5-N-acetyl-9-aminoacetamido-9-deoxy-neuraminic acid peracetylate, 5-N-acetyl-9-aminoacetamido-9-deoxy-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-aminoacetamido-9-deoxy-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-9-succinylamido-neuraminic acid, 5-N-acetyl-9-deoxy-9-succinylamido-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-9-succinylamido-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-9-succinylamido-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-9-thio-neuraminic acid, 5-N-acetyl-9-deoxy-9-thio-neuraminic acid ester peracetylate, 5-N-acetyl-9-deoxy-9-thio-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-9-thio-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-9-methylthio-neuraminic acid, 5-N-acetyl-9-deoxy-9-methylthio-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-9-methylthio-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-9-methylthio-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-9-methylsulfonyl-neuraminic acid, 5-N-acetyl-9-deoxy-9-methylsulfonyl-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-9-methylsulfonyl-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-9-methylsulfonyl-neuraminic acid ethyl ester peracetylate, 5-N-flouroacetyl-neuraminic acid peracetylate, 5-N-flouroacetyl-neuraminic acid methyl ester peracetylate, 5-N-flouroacetyl-neuraminic acid ethyl ester peracetylate, 5-N-triflouroacetyl-neuraminic acid peracetylate, 5-N-triflouroacetyl-neuraminic acid methyl ester peracetylate, 5-N-triflouroacetyl-neuraminic acid ethyl ester peracetylate, 5-N-aminoacetyl-neuraminic acid peracetylate, 5-N-aminoacetyl-neuraminic acid methyl ester peracetylate, 5-N-aminoacetyl-neuraminic acid ethyl ester peracetylate, 5-N-succinyl-neuraminic acid peracetylate, 5-N-succinyl-neuraminic acid methyl ester peracetylate, 5-N-succinyl-neuraminic acid ethyl ester peracetylate, 5-N-thioacetyl-neuraminic acid peracetylate, 5-N-thioacetyl-neuraminic acid methyl ester peracetylate, 5-N-thioacetyl-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-neuraminic acid methyl ester, 5-N-acetyl-9-deoxy-neuraminic acid ethyl ester, 5-N-acetyl-9-deoxy-neuraminic acid methyl ester peracetylate, 5-N-acetyl-9-deoxy-neuraminic acid ethyl ester peracetylate, 5-N-acetyl-9-deoxy-neuraminic acid peracetylate, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid methyl ester, 5-N-acetyl-9-deoxy-9-iodo-neuraminic acid, methyl ester peracetylate. 5. Method for the production of glycoconjugates according to claim 1, characterized in that a sialic derivative of general formula I as given in claim 2, wherein R1 represents hydrogen or lower alkyl up to 5 carbon atoms, which may be branched, unbranched, acyclic, alicyclic or cyclic, R2 is acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic, R3 is a halogen atom, a methylsulfide group, a methylsulfate group or acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic. R4 represents, independently from each other, hydrogen, acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched, unbranched, acyclic, alicyclic or cyclic, R5 is hydrogen or a halogen atom and R6 represents hydrogen, acetyl, thioacetyl or succinyl, which can be substituted with up to 3 fluoro atoms or an amino group, or an acyl or thioacyl group with up to 5 carbon atoms in the alkyl moiety, which may be branched unbranched, acyclic, alicyclic or cyclic, is incorporated into a living body, especially to a mammal or cells derived from mammals and lower eukaryotes. 6. (Canceled). 7. Use of a sialic derivative of general formula I as given in any of the claims 1 to 5, for producing a pharmaceutical composition, for immunosuppression, cell protection, stimulation of hematopoesis regulation of hormonal secretion and hormonal activation. |
Integrated multiplexer/de-multiplexer for active-matrix display/imaging arrays |
This invention presents Vt-shift invariant integrated multiplexer and de-multiplexer circuits that can be fabricated with a-Si:H, poly-crystalline silicon, or organic/polymer TFTs. The de-multiplexer and multiplexer includes a plurality of TFTs which are connected in series, and a drive TFT. These circuits are used with active matrix displays to control the gate addressing, and with imaging arrays to multiplex the read-out data. |
1. A drive circuit for driving a pixel array, the drive circuit comprising: an output terminal for driving a transistor in a pixel array; a drive transistor for transferring a gate selecting signal to the output terminal; and one or more control transistors for switching the drive transistor in response to one or more control signals, the drive transistor, the control transistors and the transistor in the pixel array being a thin film transistor. 2. The drive circuit as claimed in claim 1, wherein the control transistors are connected in series between a terminal receiving a switching signal and the gate of the drive transistor. 3. The drive circuit as claimed in claim 2 further comprising an output buffer connected to the source terminal of the drive transistor, the drain terminal of the drive transistor receiving the gate selecting signal. 4. The drive circuit as claimed in claim 2, wherein each of the control signals has a duty cycle of 50%. 5. The drive circuit as claimed in claim 3, wherein each of the control signals, the switching signal and the gate selecting signal is at a negative voltage in a logic state “low”, and each of the control signals, the switching signal and the gate selecting signal is at a positive voltage in a logic state“high”. 6. The drive circuit as claimed in claim 1, wherein the thin film transistor is derived from an inorganic or organic/polymer material. 7. The drive circuit as claimed in claim 6, wherein the thin film transistor is an amorphous silicon transistor or a polycrystalline silicon transistor. 8. (Cancelled) 9. The driver as claimed in claim 1 wherein the driver includes: a plurality of de-multiplexers, each of which drives a corresponding gate line in the pixel array; and one or more control signal lines for activating the de-multiplexers, the de-multiplexer including: the output terminal connected to the corresponding gate line in the pixel array; the drive transistor; and one or more control transistors for switching the drive transistor in response to the control signals from the control signal lines. 10. The driver as claimed in claim 9, wherein the de-multiplexer is integrated with the pixel array. 11-14. (Cancelled) 15. The driver as claimed in claim 9, wherein the control signal lines are activated such that only one de-multiplexer is activated at one time. 16-18. (Cancelled) 19. A read circuit for reading data from a data line in a pixel array, the read circuit comprising: an input terminal connected to a data line in a pixel array, data in the pixel array transferred to the data line by a transistor in the pixel array; an output terminal; a drive transistor for transferring the data to the output terminal; and one or more control transistors for switching the drive transistor in response to one or more control signals, the drive transistor, the control transistors and the transistor in the pixel array being a thin film transistor. 20. The read circuit as claimed in claim 19, wherein the control transistors are connected in series between a terminal receiving a switching signal and the gate of the drive transistor. 21. The read circuit as claimed in claim 20, wherein each of the control signals has a duty cycle of 50%. 22. The read circuit as claimed in claim 20, wherein each of the control signals and the switching signal is at a negative voltage in a logic state “low”, and is at a positive voltage in a logic state “high”. 23. The read circuit as claimed in claim 19 further comprising a pull up network circuit for pulling up a gate voltage of the drive transistor in response to a switching signal. 24-25. (Cancelled) 26. The read circuit as claimed in claim 23, wherein the control transistors are connected between the gate of the drive transistor and a ground. 27. The read circuit as claimed in claim 23 further comprising an output buffer for receiving a signal on the input terminal. 28. The read circuit as claimed in claim 19, wherein the thin film transistor is derived from an inorganic or organic/polymer material. 29. The read circuit as claimed in claim 19, wherein the thin film transistor is an amorphous silicon transistor or a poly-crystalline silicon transistor. 30. (Cancelled) 31. The read circuit as claimed in claim 19, wherein the pixel array including a plurality of data lines, each of which is connected to a transistor for transferring data to the data line, the read circuit comprising: a plurality of multiplexers, each of which is connected to a data line in a the pixel array; and one or more control signal lines for activating the multiplexers, the multiplexer including: the input terminal connected to a corresponding data line in the pixel array; the output terminal; the drive transistor; and the one or more control transistors for switching the drive transistor in response to the one or more control signals. 32. The read circuit as claimed in claim 31, wherein the multiplexer is integrated with the pixel array. 33. The read circuit as claimed in claim 31, wherein the control transistors are connected in series between a terminal receiving a switching signal and the gate of the drive transistor. 34-35 (Cancelled) 36. The read circuit as claimed in claim 31, wherein the multiplexer further comprises a pull up network circuit for pulling up a gate voltage of the drive transistor in response to a switching signal. 37-39. (Cancelled) 40. The read circuit as claimed in claim 36, wherein the multiplexer further comprises an output buffer for receiving a signal on the input terminal. 41. The read circuit as claimed in claim 31, wherein the control signal lines are activated such that only one multiplexer is activated at one time. 42. The read circuit as claimed in claim 31, wherein the thin film transistor is derived from an inorganic or organic/polymer material. 43. The read circuit as claimed in claim 31, wherein the thin film transistor is an amorphous silicon transistor or a polycrystalline silicon transistor. 44. (Cancelled) 45. The drive circuit as claimed in claim 1 further comprising a pull up network circuit for pulling up the gate line voltage and/or a pull down network circuit for pulling down the gate line voltage. 46. (Cancelled) 47. The driver as claimed in claim 9, wherein the de-multiplexer further comprising a pull up network circuit for pulling up the gate line voltage and/or a pull down network circuit for pulling down the gate line voltage. 48. (Cancelled) 49. A drive circuit for driving a pixel array, the drive circuit comprising: a pull up network circuit for pulling up a gate voltage of a switching transistor in a pixel array in response to a gate selecting signal; and a pull down network circuit for pulling down the gate voltage in response to one or more control signals; the pull down network circuit including one or more transistors, the transistors of the pull down network circuit and the switching transistor being a thin film transistor. 50. The drive circuit as claimed in claim 49, wherein the transistors of the pull down network circuit are connected between a terminal which is connected to the gate of the switching transistor and a ground. 51-52. (Cancelled) 53. The drive circuit as claimed in claim 49, wherein the control signals are at a negative voltage in a logic state “low”. 54. The drive circuit as claimed in claim 49, wherein the gate selecting signal is at a negative voltage in a logic state “low”. |
<SOH> BACKGROUND OF THE INVENTION <EOH>The most popular addressing method in large area displays is active matrix addressing where the gate and data lines form the rows and columns of the grid-like structure. FIG. 1 is a diagram showing an active matrix array 100 of an active matrix display. The active matrix array 100 has a plurality of pixels 106 . The pixels are controlled by transistors in the electronic backplane. The active matrix array has at least one transistor per pixel that acts as an analog switch. The switching transistor either enables or disables writing of data to that pixel. In FIG. 1 , Thin Film Transistor (TFT) 108 is shown as the switching transistor, which is connected to a data line 102 and a gate line 104 . The switching transistors of the display array are controlled by a de-multiplexer (also known as a gate driver). The purpose of the gate driver is to sequentially activate every row of the display while data is being written to that row. This data is stored and retained by the active pixels until they get new data in the next frame. This method of writing data to a display array is known as row-by-row addressing. Currently, amorphous silicon (a-Si:H), polycrystalline silicon, or organic/polymer materials can be used for making the switching transistors in display pixels. In a-Si:H and polycrystalline silicon, the TFTs suffer from electrical-stress induced meta-stability problems. Therefore, they are not usually used in the implementation of the driving circuitry. However, if the metastability problems can be overcome, there are significant benefits including cost savings in implementing integrated gate drivers on the display instead of having external chips. It is also desirable to provide a gate multiplexers/de-multiplexers that can also be integrated with active-matrix imaging arrays, where the imaging pixels are activated row-by-row during image read-out. |
<SOH> SUMMARY OF THE INVENTION <EOH>The objective of this invention is to provide an integrated gate de-multiplexer and read-out multiplexer that can be integrated on to a a-Si:H, poly-crystalline silicon, or organic/polymer display or imaging arrays. Further, it is an object of the present invention to provide an integrated gate de-multiplexer and read-out multiplexer that overcomes the material metastability, and has threshold voltage (Vt-shift) invariant operation over the lifetime of the array. In accordance with an aspect of the present invention, there is provided a drive circuit for driving a pixel array, which includes an output terminal for driving a transistor in a pixel array, a drive transistor for transferring a gate selecting signal to the output terminal, and one or more control transistors for switching the drive transistor in response to one or more control signals. The drive transistor, the control transistors and the transistor in the pixel array are thin film transistors. In accordance with a further aspect of the present invention, there is provided a driver for driving a pixel array. The pixel array includes a plurality of gate lines, each of which is connected to a gate of a switching transistor. The driver includes a plurality of de-multiplexers, each of which drives a corresponding gate line in a pixel array, and one or more control signal lines for activating the de-multiplexers. The de-multiplexer includes an output terminal connected to the corresponding gate line in the pixel array, a drive transistor for transferring a gate selecting signal to the output terminal, and one or more control transistors for switching the drive transistor in response to control signals from the control signal lines. The drive transistor, the control transistors and the switching transistor in the pixel array are thin film transistors. In accordance with a further aspect of the present invention, there is provided a read circuit for reading data from a data line in a pixel array. The read circuit includes an input terminal connected to a data line in a pixel array, data in the pixel array transferred to the data line by a transistor in the pixel array, an output terminal, a drive transistor for transferring the data to the output terminal and one or more control transistors for switching the drive transistor in response to one or more control signals. The drive transistor, the control transistors and the transistor in the pixel array are thin film transistors. In accordance with a further aspect of the present invention, there is provided a read circuit for reading data from an pixel array. The pixel array includes a plurality of data lines, each of which is connected to a transistor for transferring data to the data line. The read circuit includes a plurality of multiplexers, each of which is connected to a data line in a pixel array and one or more control signal lines for activating the multiplexers. The multiplexer includes an input terminal connected to a corresponding data line in the pixel array, an output terminal, a drive transistor for transferring the data to an output terminal and one or more control transistors for switching the drive transistor in response to one or more control signals. The drive transistor, the control transistors and the transistor in the pixel array are thin film transistors. In accordance with a further aspect of the present invention, there is provided a drive circuit for driving a pixel array, which includes a pull up network circuit for pulling up a gate voltage of a switching transistor in a pixel array in response to a gate selecting signal, and a pull down network circuit for pulling down the gate voltage in response to one or more control signals. The pull down network circuit includes one or more transistors. The transistors of the pull down network circuit and the switching transistor are thin film transistors. According to the invention, gate de-multiplexers and read-out multiplexers can be integrated into arrays, such as active-matrix display/imaging arrays, and the integrated gate de-multiplexers and read-out multiplexers can ensure stability of the transistor. Other aspects and features of the present invention will be readily apparent to those skilled in the art from a review of the following detailed description of preferred embodiments in conjunction with the accompanying drawings. |
Wire electrical discharge machining apparatus |
A workpiece and a wire electrode are positioned near each other with a gap therebetween. A high-frequency voltage applying unit applies a high-frequency voltage and a direct current voltage applying unit applies a direct current voltage, to the workpiece and the wire electrode. A detecting unit detects a low frequency component of a voltage occurring in the gap as a result of the electrical disch |
1. A wire elecrical discharge machining apparatus that machines a workpiece based on a high-frequency voltage applied in an inter-pole gap between a wire electrode and the workpiece, comprising: a direct current voltage applying unit that applies a direct current voltage in the inter-pole gap; and a low frequency component detecting unit that detects a low frequency component from a voltage in the inter-pole gap. 2. The wire electrical discharge machining apparatus according to claim 1, farther comprising: a displaying unit that displays a state in the inter-pole gap based on an output of the low frequency component detecting unit. 3. The wire electrical discharge machining apparatus according to claim 1, farther comprising: a feed rate controlling unit that controls a feed rate based on an output of the low frequency component detecting unit. 4. The wire electrical discharge machining apparatus according to claim 1, farther comprising: a displaying unit that displays a state in the inter-pole gap based on an output of the low frequency component detecting unit; and a feed rate controlling unit that controls a feed rate based on an output of the low frequency component detecting unit. |
<SOH> BACKGROUND ART <EOH>It is known that a finely machined surface can be obtained in a wire electrical discharge machining when an electrical discharge of short duration is generated repeatedly by applying a high-frequency voltage in a gap (hereinafter, “a machining gap” or “an inter-pole gap”) between a wire electrode and a workpiece. For example, Japanese Patent Application Laid-open No. S61-260915 (power source for electric discharge machining) discloses that a machined surface less than 1 μmRmax can be obtained if a frequency of the high-frequency voltage that is applied in the machining gap is between 1.0 megahertz and 5.0 megahertz. Japanese Patent Application Laid-open No. H7-9258 (method and device for electric discharge machining, electrostatic capacitance device and inductance varying device both applicable thereto) disclose that a machined surface less than 5 μmRmax can be obtained if the frequency of the high-frequency voltage applied in the machining gap is between 7.0 megahertz and 30 megahertz. FIG. 1 is a block diagram of a power supply source that is commonly used in a wire electrical discharge machining apparatus. This power supply source includes a direct current power source 101 and a high-frequency oscillator amplification-circuit 102 . The direct current power source 101 supplies power to the high-frequency oscillator amplification-circuit 102 . The high-frequency oscillator amplification-circuit 102 generates a high-frequency voltage based on the power received from the direct current power source 101 . The high-frequency voltage is applied to a wire electrode 103 and a workpiece 104 . The wire electrode 103 and the workpiece 104 are located, at a site where an electric discharge is to be generated, in an opposite manner with a designated gap, i.e., a machining gap 105 . The direct current power source 101 generates a constant direct current voltage or a direct current power in response to an external instruction. The high-frequency oscillator amplification-circuit 102 generates the high-frequency voltage based on the direct current voltage or the direct current power. The high-frequency oscillator amplification-circuit 102 applies a high-frequency voltage in the machining gap 105 . As a result, a high-frequency electric discharge is generated in the machining gap 105 and the workpiece is machined by the energy of the electric discharge. FIG. 2 is an example of a waveform of no-load voltage that is the high-frequency voltage applied in the machining gap. A high-frequency voltage 201 applied in the machining gap has a waveform in which the voltage oscillates symmetrically around a ground (GND) electrical potential, which is a reference level. It is known that if the high-frequency voltage 201 of a frequency more than 1 megahertz is applied repeatedly and continuously, a finely machined surface can be obtained. FIG. 3 is an example of a waveform of voltage when the high-frequency voltage is applied with a stop interval in which the voltage is not applied. FIG. 3 illustrates an example of a case when a high-frequency voltage 301 is applied repeatedly with a stop interval 302 of duration T2. A power supply source that generates this type of high-frequency voltage has been put in use. It is known that this type of high-frequency voltage improves the smoothness and finishing line of the surface of the workpiece. It is known in the art to control the machining based on a real voltage (hereinafter, “inter-pole gap voltage”) in the machining gap in order to maintain a stable machining state. This control is carried out as follows. When the electrical discharge starts between the wire electrode and the workpiece, the inter-pole gap voltage decreases. The more the wire electrode approaches the workpiece and a discharge period becomes shorter, namely the more frequently the discharge occurs, the inter-pole gap voltage decreases further. Therefore, it is possible to estimate a width of the machining gap and determine whether the width is too short or too long. Precisely, the inter-pole gap voltage is rectificated and transformed to a voltage that has one polarity. Based on this voltage, it is estimated whether a state in the inter-pole gap is an open state, a short circuit state, or a discharging state. The open state means that the electrical discharge does not take place. The short circuit state means that the wire electrode and the workpiece have short circuited. The discharging state means that the electrical discharge once occurred, but now the wire electrode and the workpiece have short circuited. A relative feed rate between the wire electrode and the workpiece is controlled (hereinafter, “an inter-pole gap servo”) based on the state determined in such a manner that the workpiece is machined in a stable manner. The high-frequency voltage that has a frequency of more than several megahertz is beyond an operating limit of a rectification circuit of the wire electrical discharge machining apparatus. As a result, it is difficult to monitor the state in the machining gap and, therefore, it is difficult to control the relative feed rate between the wire electrode and the workpiece. In other words, when a high-frequency power supply source is used, it is sometimes difficult to maintain the stable machining state. It is possible to maintain the stable machining condition when relative movement between the wire electrode and the workpiece is performed at a constant speed. An example of such machining is a machining when the machining amount changes very little, such as when performing finishing machining of a roughly machined workpiece. However, when the machining amount changes considerably as a result of change in the shape of the workpiece during machining, as it is difficult to maintain a stable machining state because it is difficult to monitor the state in the machining gap, traces of lines get easily formed on the machining surface. The shape of the workpiece may change due to distortion of the workpiece during the machining. As a result, at the time of a first-cut or if there is a variation in the machining amount, the high-frequency power supply source gives bad results. Thus, although the wire electrical discharge machining apparatus with the high-frequency power supply source gives better results in general, there are drawbacks that need to be taken care of to meet strict market demands. One approach to solve this problem is to control the relative feed rate based on a state in the inter-pole gap. For example, Japanese Patent Application Laid-open No. H10-43951 (wire electric discharge machining device) discloses a technology to detect whether the wire electrode and the workpiece have made a physical contact and then control the feed rate based on a result of the detection in a manner to achieve stable machining condition. However, in this publication does not disclose a method to detect whether the wire electrode and the workpiece have made a physical contact. The present invention has been made to solve the above problems, and it is an object of the present invention to provide a wire electrical discharge machining apparatus that can detect the state in the inter-pole gap and that can control the feed rate depending on the detected state. |
<SOH> BRIEF DESCRIPTION OF THE DRAWINGS <EOH>FIG. 1 is a general block diagram of a power supply source that is used in a wire electrical discharge machining apparatus; FIG. 2 is an example of a waveform of no-load voltage that is a high-frequency voltage applied in a machining gap; FIG. 3 is an example of a waveform of voltage when the high-frequency voltage is applied repeatedly with a stop interval in between; FIG. 4 is a concrete circuit diagram of the wire electrical discharge machining apparatus according to a first embodiment of the present invention; FIG. 5 is a waveform diagram to explain an inter-pole gap voltage; FIG. 6 is a illustration of a principle to detect a direct current voltage component from the voltage in the inter-pole gap; and FIG. 7 is a concrete block diagram of a wire electrical discharge machining apparatus according to a second embodiment of the present invention. detailed-description description="Detailed Description" end="lead"? |
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