The present invention relates to methods to control the spread of recombinant DNA molecules between sexually compatible plants of differing genetic composition. The invention describes the production of transgenic plants that comprise recombinant traits of interest or concern linked to repressible lethal genes. The lethal genes are blocked by the action of repressor molecules produced by the expression of repressor genes located at a different genetic locus. The lethal phenotype is only expressed after the segregation of the repressible lethal gene construct and the repressor gene following meiosis. The present invention may be employed for both open-pollinated and hybrid seed production systems and may be used to maintain genetic purity by blocking unintended introgression of genes from plants devoid of the specific repressor gene. The invention includes methods that impart traits that are desirable for environmentally responsible heterologous protein production, to genetic material used to impart said traits and to new plants and products derived by said methods.

The increasing number and diversity of plants containing novel traits derived from recombinant DNA research present both environmental and commercial concerns. The concerns arise from the potential for novel traits to spread by pollen to sexually compatible plants in a natural or cultivated population.
Plants with new and altered traits imparted by genetic technologies and recombinant DNA technology in particular are now viewed as the cornerstone of the crop biotechnology industry. Currently a considerable number of crops plants with novel traits that originated from tissue culture, somatoclonal variation or mutation as well as genetic engineering are undergoing field trials and the first stages of commercial release. These plants not only include conventional crops grown on an annual basis, but other plants such as trees or shrubs which comprise novel traits and are perennial in nature.
Modern crop varieties comprise both individual genes that confer a particular trait and combination of genes assembled through conventional plant breeding. Accordingly, as more novel traits are developed and incorporated into modern crop varieties, it is valuable to have a means to preserve genetic compositions, including those of specific crop varieties, cultivars or breeding lines. Of particular value is the preservation of crops which carry traits not usually found in the crop; for example, plants which produce novel oil, meal or other components or those plants modified to produce speciality chemicals. Additionally, perennial plants such as trees are being produced which carry novel traits such as altered lignin levels, insect and fungal resistance and herbicide tolerance.
Novel traits are introduced into plants by conventional breeding or genetic engineering. However, to date neither route provides features that can be routinely used for maintaining germplasm purity, or controlling persistence or potential spread of the novel trait. Current vectors and genetic compositions typically do not address two important issues: (1) commercial issues such as the prevention of transformed crop plants or elite varieties from contaminating other commercial productions, or the prevention of introgression of alien germplasm from closely related cultivars or plant species, and; (2) environmental issues such as the removal of transformed crop plants or related species that have acquired the genes in question from non-agricultural environments. Additionally current transformation methods do not provide the means for reducing the introduction of genes via pollen mediated out-crossing to other cultivars or related species (either wild or cultivated).
The single largest immediate risk for the use of many crops with novel traits is the risk of contamination among commercial productions of the same crop species. The risk of a crop species such as oilseed rape or canola (Brassica napus) to become a weed or to cross with wild weedy relatives is modest compared with the near certainty of crossing with other commercial productions of canola, especially where large production areas exist. In the past this has not been a significant problem for farmers and commercial processors for several reasons. First, breeding objectives have been relatively uniform for canola crop; second, only a small number of cultivars have comprised 90-100% of the total acreage grown by farmers; and third, the only speciality type, traditionally cultivated, high erucic acid industrial oil cultivars have been grown in physical isolation. Accordingly, cross contamination of food quality canola varieties with genes conferring high erucic acid has not been a serious issue.
Recently additional unique varieties have been released. These include varieties that carry recombinant genes which confer tolerance to herbicides and varieties developed by conventional breeding which have variations in fatty acid profile, such as high oleic acid. Purity of seed, both during production and harvesting of canola seed for crushing and processing is now a growing issue. Because of the impending modification of canola with numerous additional recombinant genes that impart different properties to the oil (e.g. high laurate content) or the use of plants as producers of heterologous proteins such as pharmaceuticals, potentially serious industrial cross contamination may be anticipated.
These issues extend to many crops in addition to Brassica oilseeds. In maize, increasing emphasis on herbicide tolerance, insect resistance and diversification of modified end products (eg. starch, oil, meal) clearly indicates that many different traits will be incorporated in the corn crop. As some maize varieties are destined for specialized use, such as wet milling or feed, or even production of pharmacologically important proteins, the issue of segregation of these speciality types from the mainstream is relevant. Considering that corn pollen can sometimes travel significant distances, a genetic means to control pollination is be highly advantageous.
Similarly, the proximity of perennial plants to their wild relatives is a problem. For instance, a transgenic tree expressing insect tolerance could cross with a wild species of tree to create a hybrid that expresses insect tolerance. Under managed conditions such as plantations, insect resistance would not have a significant environmental impact. However, should the insect resistance trait become widespread in a natural forest population a serious ecological problem could result. Insect populations are part of the food chain in a forest system and reduced levels of insects could lead to a collapse of the predator population, which is often native bird species. Accordingly, for unmanaged systems control of the spread of genes that may carry environmental consequences is a highly desirable goal.
Currently physical isolation combined with border rows that function as pollen traps have been employed to contain transgenic plants under study and development. This method, however, is impractical for widespread cultivation. Moreover, with increasing production and distribution of an increasing number of different transgenic types, the potential for contamination increases dramatically. This issue has recently become a major concern for the oilseed rape industry and will become a greater issue for other major crops (eg. corn) as the numbers of different recombinant and speciality genotypes reach the market place.
In addition to cross-contamination among commercial crop productions, another concern is the potential spread of crops used as vehicles for producing heterologous proteins of commercial or medicinal value. These novel protein products can potentially contaminate plants destined for food use and export. Although production standards can be implemented that will attempt to preserve the identity of individual transgenic lines and reduce unintended contaminations, the outflow of genes to other cultivars will eventually occur. The potential spread of genes that cannot be easily identified, e.g. by herbicide tolerance, nor impart a distinctive morphology has yet to be addressed by government or industry.
Methods which control the spread of transgenes into the environment or other commercial cultivars are also useful for preventing the introgression of alien germplasm into identity-preserved commercial varieties. In this regard xe2x80x9calien germplasmxe2x80x9d is defined as any germplasm which does not comprise the full complement of traits of the identity-preserved cultivar. Accordingly alien germplasm can include both sexually compatible wild relatives and other commercial varieties of the crop. With an increasing number of plants carrying novel traits being contemplated for commercial production, methods that prevent the contamination of both seed production and commodity production will provide a valuable means to maintain germplasm purity and identity preservation.
As an example, many enzymes have been tested that alter plant oil production in oilseed crops such as soybean corn and canola. The same plant species have been used for producing inedible short chain or long chain industrially fatty acids as well as edible oil. Since modified oil seeds must be isolated to ensure pollen carrying the oil modification genes does not contaminate edible oil variety seeds, this poses a growing problem for the seed production industry. The isolation distances routinely practiced in seed production for many crops may not be sufficient to ensure required levels of purity. Where crop plants are used to produce speciality products such as pharmaceutically active compounds, even minor contamination of germplasm is highly undesirable.
Oil seed crops such as canola typically shatter seed before harvest. This results in significant numbers of volunteer plants in subsequent years, potentially contaminating subsequent commercial productions both by crossing and by direct effects of the pollen on developing grain (xenia effects). In addition, seeds retained and distributed by farmers for future planting could contribute to contamination problems.
For perennial plants, the long life of trees and the presence of indigenous wild relatives raise additional concerns. Some trees take many years to flower, producing enormous amounts of pollen that can last for many years and are especially suited for widespread wind pollination. Transgenic trees therefore pose special problems and may require mechanisms to control gene flow to wild relatives.
It has been suggested that some new crop types, through hybridization with wild relatives, may invade natural ecosystems. This and related issues have been extensively debated (eg. University of California, Risk assessment in agricultural biology: proceedings of an international conference, 1990, Casper, R., and Landsman, J., 1992, The bio-safety results of field tests of genetically modified plants and microorganisms. Proceedings of the 2nd International Symposium on The Biosafety Results of Field Tests of Genetically Modified Plants and Microorganisms, 1992 Goslar, Germany, Dale, P. et al., 1992, The field release of transgenic plants. The British Crop Protection Council. Brighton Crop Protection Conference: Pests and Diseases, Vols. I, II and III., Proceedings of the 3rd International Symposium on The BioSafety Results of Field Tests of Genetically Modified Plants and Microorganisms, 1994, Monterey, Calif., D. D. Jones, 1994).
The consensus of these studies and experimental results achieved to date support the view that the degree of potential spread of transgenes to wild relatives is highly dependent upon the species and environmental conditions. Crossing with relatives is not likely with some species and probable for others (Raybould and Grey, J. Applied Ecology 30: 199-219, 1993). Many crops are highly specialized and adapted to non-competitive cultivation practices and thus are not generally considered a serious environmental risk on their own (Dale et al., Plant Breeding 111:1-22, 1993, Fishlock, D., The PROSAMO Report, published by the Laboratory of the Government Chemist, Queens Road, Teddington, Middlesex, UK TW11 0LY). The potential for environmental problems due to, for example, the inclusion of a virus coat protein gene that has potential for viral recombination and the evolution of new viruses with an extended host range, is currently unknown (Gal S., et al., Virology 187:525-533, Grimsley, N., et al., EMBO Journal 5: 641-646, 1986, Lecoq, H., et al., Molec. Plant Microbe Interact. 6:403-406, 1993. Tepfer, M., Biotechnology 11: 1125-1132, 1993). Accordingly there is a need for methods to restrict the potential flow of this type of genes or to selectively eliminate those plants which contain such genes.
Attempts have been made to develop methods to specifically remove or identify plants that contain novel traits introduced by recombinant DNA. For example, the use of a conditionally lethal gene, i.e. one which results in plant cell death under certain conditions, has been suggested as a means to selectively kill plant cells containing a specific recombinant DNA. Recently the development of genes which are conditionally lethal in plants have been described (eg WO 94/03619). However, methods using these genes have been restricted to the application of a substance that triggers the expression of the lethal phenotype. For widespread agricultural practices, these methods have serious limitations.
An example of a conditionally lethal gene is the Agrobacterium Ti plasmid-derived oncogene commonly referred to as xe2x80x9cgene 2xe2x80x9d or xe2x80x9concogene 2xe2x80x9d. The gene encodes the enzyme indole acetamide hydrolase (IAMH) that hydrolyzes indole acetamide, a compound that has essentially no phytohormone activity, to form the active auxin phytohormone indole acetic acid. The enzyme IAMH is capable of hydrolyzing a number of indole amide substrates including naphthalene acetamide, resulting in the production of the well known synthetic plant growth regulator naphthalene acetic acid (NAA). Use of the IAMH gene for roguing plants has been described by Jorgenson (U.S. Pat. No. 5,180,873). The method requires application of NAM to discriminate plants which carry the conditionally lethal gene.
Other enzymes may also be used as conditionally lethal genes. These include enzymes which act directly to convert a non-toxic substance to a toxin, such as the enzyme methoxinine dehydrogenase, which converts non-toxic 2-amino-4-methoxy-butanoic acid (methoxinine) to toxic methoxyvinyl glycine (Margraff, R., et al., 1980, Experimentia 36: 846), the enzyme rhizobitoxine synthase, which converts non-toxic 2-amino-4-methoxy-butanoic acid to toxic 2-amino-4-[2-amino-3-hydroxypropyl]-trans-3-butanoic acid (rhizobitoxine) (Owens, L. D, et al., 1973, Weed Science 21: 63-66), the de-acylase enzyme which acts specificlly to convert the inactive herbicide derivative L-N-acetyl-phosphinothricin to the active phytotoxic agent phosphinothricin (Bartsch, K. and Schultz, A., EP 617121), and the enzyme phosphonate monoester hydrolase which can hydrolyze inactive ester derivatives of the herbicide glyphosate to form the active herbicide (Dotson S. B., and Kishore G. M., 1993, U.S. Pat. No. 5,254,801). Other conditionally lethal genes may be engineered from lethal genes. A lethal gene which is expressed only in response to environmental or physiological conditions is lethal under those conditions. For example, a gene that encodes a lethal activity may be placed under the control of a promoter that is induced in response to a specific chemical trigger or an artificial or naturally occurring physiological stress. In this fashion the expression of the lethal gene activity is conditional on the presence of the inducer.
The expression of the conditionally lethal gene that acts on a non-toxic substance to convert said substance to a toxic substance is typically regulated by a promoter that is a constitutive promoter expressed in all or most cell types or a developmentally regulated promoter expressed in certain cell types or at certain stages of development. Any promoter that provides sufficient level of expression can be used. However, in practice promoters that provide high levels of expression for extended periods offer the best opportunities to remove unwanted plants.
The need to apply a chemical to induce the lethal phenotype reduces the utility of a conditionally lethal gene. The widespread application of chemicals may be impractical and raise additional environmental concerns. Accordingly the use of conditionally lethal genes as currently described is not ideally suited for general applications since intervention is required to express the lethal phenotype.
The possibility of using a repressed lethal gene to limit the persistence of hybrid crops has been suggested recently by Oliver et al (patent application WO 96/04393). In this system expression of a lethal gene is blocked by a genetic element that binds a specific repressor protein. The repressor protein is the product of a repressor gene typically of bacterial origin. The genetic element that binds the repressor protein is referred to as a blocking sequence and is constructed such that it further comprises DNA sequences recognized by a DNA recombinase enzyme (e.g. the CRE enzyme). Plants that contain said blocked lethal gene are hybridized with plants comprising the DNA recombinase gene. Either the lethal gene or the recombinase enzyme (or both) is under control of regulatory elements that allow expression only at a specific stage of plant development (e.g. seed embryo). Consequently, the recombinase function in the resulting F1 hybrid plant removes the specific blocking sequence and activates the lethal gene so that no F2 plant is produced. Notably, this scheme cannot control outcrossing of germplasm that carries the novel trait nor introgression of alien germplasm. The method does not apply to self- or open-pollinating varieties. Accordingly, the method is useful only as a means to restrict use (e.g. re-planting) to F1 hybrid seed.
Methods to eliminate recombinant DNA sequences used to obtain transformants such as selectable markers have been developed. Use of a transposase or recombinase to remove selected recombinant sequences from transgenic crop plants has been described in U.S. Pat. No. 5,482,852 (Biologically Safe Transformation System, by Yoder and Lassner). This invention describes a method for removing vector and marker gene sequences by enclosing them within a transposon. The sequences are subsequently removed by crossing the plant to a plant with transposase function.
No published method, however, addresses the problem of contamination of related varieties by cross pollination. The art also does not provide a means to prevent the introgression of alien germplasm by pollination with related pollen, even pollen from the same variety but lacking the genetic trait(s).
Therefore, a method that limits outcrossing and introgression without intervention is needed for management and control of novel traits and crops with novel traits. A mechanism to control cross-contaminations among commercial crops is also needed. Such a mechanism is also needed in the management of perennial crops such as trees, shrubs and grapevines. In particular any mechanism which does not require intervention in order to function is ideally suited for perennial crops. The present invention describes methods and genetic compositions which respond to these needs.
The present invention comprises methods and recombinant DNA compositions that block the spread and persistence of genes in other cultivars of the same species or related species, resulting from unintended outcrossing by pollen produced by plants containing said recombinant DNA. The invention further ensures that introgression of alien germplasm is eliminated in a selfing population.
The present invention relates to novel recombinant DNA constructs that impart a novel feature to plants containing the recombinant DNA. This feature permits viable seed to be formed only on plants that contain the full complement of the recombinant DNA. The present invention further provides a means to ensure the sexual isolation of germplasm or genetic traits within a defined population through the expression of a trait that is lethal in plants which do not comprise the full complement of the recombinant DNA. The invention ensures that those plants which are fertilized by the transgenic plant but which do not carry the recombinant DNA are unable to form viable seed.
The novel genetic constructs impart no morphologically obvious or easily detectable phenotype to plants. They comprise silent genes that are expressed only when an unintended sexual cross occurs. An unintended cross results in expression of a lethal trait and the undesired plant cells are eliminated. Accordingly the invention restricts the formation of viable seed via outcrossing with sexually compatible species. The novel DNA constructs further provide a means to effectively reduce the introgression of traits from cross-pollination with pollen from sexually compatible species that lack the constructs.
The present invention provides a genetic trait encoded within DNA constructs that ensures that specific cultivars or breeding lines are not contaminated with alien germplasm or contaminate other cultivars and breeding lines. This provides a convenient means to genetically isolate the transgenic plant. The novel DNA constructs may be used as a means to ensure germplasm purity during seed production and the production of the commodity in the field and can be used in both open pollinated and hybrid crop varieties.
Linkage of the novel DNA constructs to DNA molecules that encode novel agronomic or phenotypic traits ensures that the novel agronomic or phenotypic trait does not persist outside of the genotype into which it was introduced. This aspect of the invention is useful in the management of crops with novel agronomic or phenotypic traits or crops with unique combinations of conventional traits developed through plant breeding techniques.
In one embodiment, the invention provides a genetic system comprising two DNA constructs. One DNA construct comprises a dominant repressible lethal gene that, when active, results in cellular death, and whose expression is inhibited in plant cells which contain a second DNA construct comprising a repressor gene, the repressor gene being located at a locus that segregates independently from the repressible dominant lethal gene. The repressor gene encodes a repressor molecule which may be a DNA binding protein, a direct inhibitor of the lethal gene activity, or an RNA, ribozyme or antisense RNA capable of inhibiting the lethal phenotype.
FIG. 1 illustrates the genetic constructs that may be employed in this embodiment of the invention
In a preferred embodiment, the dominant repressible lethal gene is under the control of a seed specific promoter and the gene encoding a repressor molecule is located at a locus that segregates independently from the repressible dominant lethal gene. Both the repressible dominant lethal gene and the repressor gene are in the homozygous state. Self pollination maintains this genetic combination.
In another preferred embodiment, the DNA construct further comprises a conditionally lethal gene linked to the repressible lethal gene. The conditionally lethal gene can be activated by the application of a chemical or physiological stress, ensuring a means to completely eliminate the plants or cells containing the recombinant DNA from the environment when required. Accordingly, even self-pollinated cells containing a repressible lethal gene can be selectively removed from a population by virtue of the conditionally lethal gene.
In an additional preferred embodiment, the repressible lethal gene linked to a conditionally lethal gene is linked additionally to a gene encoding a novel trait. A second DNA construct comprises a gene encoding a repressor capable of blocking the activity of the repressible lethal gene. The separate DNA constructs are introduced into the same cells. Linkage of the novel trait to the repressible lethal gene ensures that the novel trait can not persist in related species by transfer through sexual crossing.
In a still further embodiment, the DNA constructs comprising the repressible lethal gene and the repressor gene are within a single recombinant DNA molecule which is introduced into the plant cell. The single recombinant DNA molecule further contains sequences recognized by a site specific recombinase or transposase. Recombinase or transposase activity results in the removal of the repressor gene from the inserted recombinant DNA. As an element of this embodiment, the repressor gene is reintegrated to an independently segregating locus; in particular, to the same locus on the opposite chromosome of a homologous chromosome pair. The DNA constructs that may be employed within the scope of this embodiment are illustrated in FIG. 2.
In another preferred embodiment, DNA constructs are introduced into a plant cell, comprising two repressible lethal genes and two functionally distinct repressors for the repressible lethal genes. The genes are preferably arranged so that the first repressible lethal gene is linked to the repressor capable of repressing the second repressible lethal gene, and the second repressible lethal gene is linked to the repressor capable of repressing the first repressible gene, as illustrated in FIG. 3. optionally, the constructs comprise a single recombinant DNA molecule which is introduced into the plant cell. The single recombinant DNA molecule contains sequences recognized by a site specific recombinase or transposase, whose activity results in the removal of the first repressible lethal gene and the second repressor from the recombinant DNA. As an element of this optional embodiment, plants are selected wherein the first repressible lethal gene and the linked second repressor gene are reintegrated to an independently segregating locus.
The foregoing embodiments rely on random insertion of the DNA constructs to loci that segregate independently. However, for some applications a means to introduce the recombinant DNA to a specific locus may be desirable. Accordingly, the present invention provides methods to target the recombinant DNA to a specific locus.
The use of a site specific recombinase to introduce recombinant DNA to a locus previously established in the plant genome is contemplated. A recombinase target DNA sequence recognized by a site specific recombinase is inserted into the plant genome by standard transformation procedures. The plant is made homozygous for the target DNA sequence by known methods such as selfing and selection or anther or isolated microspore culture. Alternatively a plant homozygous for said inserted sequence can be made directly by transformation of haploid cells or tissue, followed by chromosome doubling.
The appropriate recombinase expressible in plant is inserted by any of several methods such as transformation, microinjection, electroportation, etc. into plant cells homozygous for the target DNA sequence. The plant cells are then independently re-transformed with DNA constructs comprising either the repressible lethal gene or the repressor gene. These DNA constructs have been modified to include site specific recombinase recognition sequences such that the DNA construct can be inserted into the pre-existing recombinase target DNA sequence. Accordingly, plant lines are recovered that contain either the DNA construct comprising the first repressible lethal gene or the first repressor gene. By crossing said lines, plants may be recovered that contain both introduced DNA constructs (repressible lethal gene and repressor) at the same genetic locus on opposite chromosomes of a homologous chromosome pair.
Accordingly the site-specific insertion method comprises preparation of DNA constructs comprising a repressible lethal gene and in some embodiments a dominant conditionally lethal gene. The method also comprises preparation of a repressor gene which can be inserted concomitantly or independently of the lethal gene. The repressible lethal gene is repressed by the repressor encoded by the repressor gene, conveniently located at a chromosomal site that segregates independently of the inserted repressible lethal gene. It is within the scope of the present method to employ site-specific recombination as a means to target repressor and repressible lethal genes to specific sites within the plant genome, in particular to those sites at which specific recombinase recognition sites have been inserted. An illustration of the DNA constructs and steps that may be employed in this embodiment of the invention are shown in FIG. 4.
The invention provides methods and compositions that allow the genetic purity of transgenic plants to be maintained by simple self pollination in open pollinated crops. No intervention is required. The invention further provides methods for the convenient preparation, propagation and husbandry of plants containing the recombinant DNA. Genetic compositions are provided for use in open pollinated and hybrid plant production systems. Illustration of the utility of the method as employed with open pollinated crops such as Brassica napus oilseed is shown in FIG. 5, illustration of the utility of the method as employed with hybrid crops such as maize is shown in FIG. 6.
During the production of pollen, the repressible lethal gene is segregated from the repressor gene, in accordance with the genetic schemes described above. Subsequently any out-crossed plants (i.e. those plants that have inadvertently received pollen that carries the repressible lethal gene) cannot form viable seed because the newly formed seed contains no repressor to repress expression of the lethal gene. The lethal gene is repressed in selfed plants because these plants retain both lethal and repressor genes. For those embodiments which further comprise a conditionally lethal gene linked to the repressible lethal gene, plants containing these genes can be eliminated by application of a chemical or physiological stress to activate the conditionally lethal gene.
The present invention provides methods and compositions for the production of recombinant plants with substantially reduced or zero risk of gene transfer via crossing. In some embodiments, the plants can be safely and specifically removed from the growing site by application of an inexpensive and environmentally benign chemical.
The invention is well suited to the production of crop plants for large scale agricultural and industrial applications where the potential contamination of other commercial productions of the same species, via cross pollination or volunteer seed, is to be avoided. The invention further provides a mechanism of safe use and environmental protection for recombinant plants that may cause environmental damage by invasion of other habitats or that may spread their transgenes by crossing by crossing with wild weedy relatives.
The present invention provides specifically a method for producing crop plants as heterologous protein producers, without risk of contaminating other commercial productions of the same species.
The invention further provides a means to control the introgression of alien germplasm into commercial plant varieties and to maintain genetic purity of lines comprising the introduced genes. It is noted that the DNA constructs comprising these genes can be used with or without being linked to a novel trait gene to provide a means of ensuring genetic purity during seed production or production of the commodity.
For some crops, such as self-incompatible crops, the invention improves hybrid seed production via self-incompatibility. In this particular embodiment of the invention, a self-incompatible female parent is modified to carry the repressible lethal gene but not the repressor gene. The female line is unable to form viable seed. Crossing this self-incompatible female parent with pollen that carries a repressor gene results in the production of viable hybrid seed that carries both the repressible lethal gene and the repressor gene. Linkage of a novel trait such as insect resistance to the repressible lethal gene would further prevent the dissemination and persistence of the trait in related species.
The use of repressible lethal genes in self-incompatible crops eliminates the problems of breakdown of self-incompatibility in the female parent often seen in commercial seed production. This breakdown problem leads to self-seed contamination of the hybrid seed. By using repressible lethal genes, self-seed is not possible on the female parent since it lacks the repressor and is self-incompatible. A convenient means to maintain the female line (such as use of a repressor inducible under certain conditions) can be employed to increase the number of female parents. Alternatively, the line can be clonally propagated. Current mechanisms to overcome self-incompatibility include elevated carbon dioxide and other stress treatments. It is within the scope of the invention to use promoters that are inducible under the same conditions as those used to overcome self-incompatibility, as this provides a particularly convenient means to increase seed production of the female parent. The method is particularly useful for production of Brassica vegetable crops where self incompatibility is commonly applied.
The following terms are defined and used within the scope of this invention.
Alien germplasm: a gene or combination of genes or genetic traits which is not part of the specific genetic makeup of an individual crop plant or variety.
Blocking or xe2x80x9cblocksxe2x80x9d: the inhibition of a lethal gene activity by a repressor; blocking can include: the prevention of RNA transcription by binding of a repressor to a specific DNA sequence, binding of an antisense RNA or ribozyme to a primary RNA or mRNA transcript, binding of an inhibiting factor to a lethal gene product such as a RNAse or protease inhibitor binding to a toxic ribonuclease or toxic protease. Any method which prevents the expression of a lethal phenotype can be considered as xe2x80x9cblockingxe2x80x9d the lethal phenotype.
Conditionally lethal gene: a gene which confers on a plant cell a phenotype which renders the plant cell sensitive to an agent, said agent may be genetic or chemical in nature, said sensitivity ultimately leading the death of the plant cell.
Constitutive promoter: a DNA sequence capable of causing gene expression in substantially all plant cells, tissues and organs.
De-repressed lethal gene: a lethal gene that expresses the lethal phenotype due to the absence of a functional repressor.
Gene: a DNA expression cassette comprising a transcribed region under the control of a promoter further comprising a transcription termination signal.
Inducible promoter: a DNA sequence capable of causing gene expression in response to a chemical, physical or environmental inducer.
Introgression: the undesired movement of a gene or genes through sexual crossing, usually by pollen, from a plant which is not intended to be the pollen donor for the formation of seed.
Lethal gene: a gene, that when expressed in a plant cell ultimately leads to the death of the plant cell.
Lethal gene activity: a genetic activity that leads to plant cell death. A lethal gene activity can be due to a single gene or can also be the result of the combined expression of more than one gene.
Oncogene: a gene encoding an enzyme involved in tumor formation or abnormal plant growth as a result of infection of susceptible plants by Agrobacterium sp. Known oncogenes include those comprising the tmr and tms loci of the T-DNA region of the Ti plasmid.
Outcrossing: the movement of pollen from a plant of one genetic type to a sexually compatible plant of a different genetic type. Outcrossing is generally used to describe the unintended movement of pollen; however in some plant species, particularly those which are self-incompatible, outcrossing is also used to represent the normal pollination events within a population of incompatible plants.
Promoter: a DNA sequence capable of causing gene expression in a plant cell.
Repressor: a gene product that can specifically block the activity of a gene product or expression of a gene. A repressor can be a protein, RNA or a specific substance produced by the activity of a repressor gene product.
Repressor binding site: a DNA sequence that is recognized specifically by a repressor, said recognition leading to the inhibition of expression of a gene containing said repressor binding site. In some embodiments a repressor binding site may also represent a RNA sequence which is recognized by a ribozyme or antisense RNA.
Repressor gene: a DNA expression cassette capable of expressing a functional repressor.
Repressed lethal gene: a lethal gene where the lethal phenotype that is a result of the gene activity is blocked by the presence of a repressor.
Repressible lethal gene: a lethal gene, the expression of which can be inhibited by the action of a specific repressor molecule.
Responsive to a repressor: a lethal gene or lethal gene product the lethal activity of which is inhibited in response to the presence of a repressor of lethal gene activity.
Selfing: self-pollination leading to the formation of a seed or reproductive structure.
Tissue specific promoter: a DNA sequence capable of causing substantial gene expression only in a specific plant cell, organ or tissue.
Transcribed region: a DNA sequence that is transcribed under the control of a promoter. Said DNA sequence may encode a RNA capable of being translated into a protein or may encode a RNA that can specifically inhibit or prevent the expression of a gene.
Transcription termination sequence: a DNA sequence that defines the termination of transcription.

Methods and genetic compositions to limit outcrossing and undesired gene flow in crop plants


In an embodiment, control is performed to operate a three-way valve in accordance with a reaching ratio of pH adjustment. When process of certain steps is performed, bubbles of ammonia generated in a nebulizer can be extruded by air so as to be injected from an injection port together with aqua ammonia remaining in the nebulizer. Therefore, the pH adjustment can be performed while a large amount of bubbles of ammonia are prevented from being generated in the nebulizer.

Analysis of trace elements including heavy metals has been performed for material circulation research in the ocean, water quality testing of tap water, water quality testing of lake water and river water, and the like. At the time of analysis of trace elements, for the purpose of separating the trace elements from interference components or for the purpose of improving the sensitivity of analysis, there may be required the separation and concentration of the trace elements based on solid-phase extraction using a chelating resin, and the like. When the separation and concentration are performed, it is required to adjust in advance a liquid sample to optimum pH conditions.
Further, in view of the purposes of analysis of trace elements, it is desirable that contamination factors in the pH adjustment are removed as much as possible. In this regard, the present inventors have developed an indirect measurement method using color change characteristics of a pH indicator (Non Patent Literature 1). In the indirect measurement method, an optical sensor is used as a pH measuring instrument, and hence contamination due to contact between the measuring instrument and the sample can be eliminated. Further, in the indirect measurement method, methyl yellow, methyl orange or methyl red is used as the pH indicator, and ammonia is used as a pH adjusting liquid. In the indicator and the adjusting liquid which are used for the indirect measurement method, contamination due to the indicator and the adjusting liquid can be eliminated by using a high purity reagent in which the amount of metal components is suppressed to be sufficiently small.

NATIONAL INSTITUTE OF ADVANCED INDUSTRIAL SCIENCE AND TECHNOLOGY


The present invention is an exercise machine for exercising latissimus dorsi muscles, chest muscles and other muscles. The invention includes moveable arms so that the distance between the arms can be varied to accommodate the size of a user or to change the type of exercise.

1. Field of the Invention
The present invention relates to an exercise machine that exercises latissimus dorsi muscles and other muscles. In practice the present invention relates to an exercise machine having movable arms for adjusting the distance between the two arms for a user's size or for the type of exercise to be performed.
2. The Prior Art
The prior art discloses that cable or belt functioning pulldown style machines utilize a single extending and immovable arm from which a bar of varying lengths is suspended from the cable or belt. The user is forced to adjust for shoulder width and type of exercise by moving his hands along said bar. This being the case, even while facing a mirror the tendency may exist to twist the bar, favoring one's stronger side and resulting in asymmetrical muscle development. It is, however, desirable to provide a machine where two arms are utilized that are movable and can be adjusted for both the size of the user as well as to vary the type of exercise to be performed.

Exercise machine for exercising latissimus dorsi muscles and other muscles


A semiconductor device according to an embodiment, includes a plurality of gate structures; a first dielectric film; and a second dielectric film. The first dielectric film crosslinks adjacent gate structures of the plurality of gate structures so as to form a cavity each above and below in a position between the adjacent gate structures. The second dielectric film is formed as if to cover the cavity above the first dielectric film between the adjacent gate structures.

In development of a semiconductor device, particularly a semiconductor storage device, ever finer patterning is developed for memory cells to achieve larger capacities and lower costs. In a semiconductor storage device mounted with a floating gate structure such as a NAND flash memory device, the wire pitch between word lines to be a control gate in a gate portion is made ever finer. Such finer patterning of LSIs is actively promoted to achieve performance improvement such as a faster operation and lower power consumption of elements due to higher integration and the reduction in manufacturing costs. In recent years, flash memories in minimum processing dimensions of, for example, 20 nm or so have been in mass production and still finer patterning and increasing technical difficulty are expected in the future. To realize a high-quality, high-performance, and low-cost semiconductor device in the face of rapid development of ever finer patterning, it is necessary to further reduce the parasitic capacitance between gate structures. In a next-generation flash memory device, both the cell width and the width between cells are extremely narrow due to still finer patterning and a technology to provide a cavity region is under development, instead of embedding a dielectric film between gate structures in a memory cell region, to inhibit electric interference between cells. By intentionally forming a cavity instead of the dielectric film embedded between gate structures in the memory cell region, a parasitic capacitance generated between gate structures can be reduced so that performance of a memory device can be improved.
However, if a cavity is formed between gate structures, mechanical strength of the gate structure is extremely weakened and with the development of ever finer patterning of devices, a problem of cracks in the dielectric film deposited in the memory cell region and a problem of a toppling gate structure in the memory cell portion are caused.

Semiconductor device and method for fabricating semiconductor device


A contoured insole especially advantageous for users with medium to high arches is disclosed. It comprises a generally foot-shaped base extending from a heel end to a toe end, which comprises a top surface and a bottom surface. The bottom surface of the base preferably further comprises two indentations formed integrally therein in a forefoot area and a heel area. A forefoot pad and a heel pad are secured to each of said indentations. Preferably, the base is made from a polyurethane foam. The pads are made from rubber or synthetic rubber. A top sheet is coextensive with and secured to the top surface of the base. The top sheet is generally a fabric which preferably has antimicrobial characteristics. In use, the foot of the wearer, with or without a sock or stocking thereon, rests upon the top sheet in the foot-receiving compartment of a user's shoe.

Insoles are inserted in the shoes of a user to provide one or more advantages to to the comfort of the wearer or the support of the foot. Insoles are generally sold in pairs and one of each pair is adapted for use in a right shoe and the other adapted for use in a left shoe of a user. It is advantageous to provide appropriate structure to an insole so that it serves the purposes of the user.


A semiconductor device with a copper line comprises a lower portion of a copper pattern buried in an interlayer insulating film, an upper portion of the copper disposed over the upper portion of the lower copper pattern, and an upper barrier metal layer disposed over upper and side surfaces of the upper copper pattern. As a result, the copper pattern is protected by the barrier metal layers, providing a metal line with a stable structure.

1. Technical Field
The present invention relates generally to a semiconductor device and a method for manufacturing the same, and more particularly to a semiconductor device including a metal line with a stable structure.
2. Related Art
In semiconductor devices, metal lines are formed in order to electrically couple devices or other conductive lines, and contact plugs are formed in order to connect lower metal lines to upper metal lines. However, the increasing degree of integration of semiconductor devices has led to a reduction in design rules. As the design rule decreases, the area available for contact holes is diminishing, so the aspect ratio of contact holes where the contact plug is formed is gradually increasing. As a result, the difficulty and importance of processes for forming the metal line and the contact plug have received an increased amount of attention.
For the metal line, aluminum (Al) and tungsten (W), each of which has excellent electric conductivity, have been widely used. Recent research has focused on using copper (Cu) as a next generation metal line material. Copper can overcome problems associated with resistor capacitor (RC) signal delay in a highly integrated high-speed operation device because copper has a relatively high electrical conductivity and low resistance. However, because it is not easy to perform a dry etching process on copper lines, a damascene process is used in order to form copper metal lines.
A method for forming a metal line with a damascene process may be performed according to the following steps. First, an interlayer insulating film is formed over a semiconductor substrate, and the interlayer insulating film is etched to form a trench. After a barrier metal layer is formed on the surface of the trench, a copper material is deposited in the trench over the barrier metal layer. Next, a chemical mechanical polishing (CMP) process is performed to form a separated copper pattern. A protective film is then formed over the pattern to prevent the upper portion of the copper pattern to physically stabilize the pattern.
In the above-described conventional semiconductor device and manufacturing method, it is difficult to form barrier metal layers over the copper pattern after separating each copper pattern by the CMP process. Although the upper portion of the copper pattern is protected by the protective film, the upper portion of the copper pattern is exposed to atmosphere during a subsequent process for forming a contact plug.

Semiconductor device with a copper line and method for manufacturing the same


Disclosed are peptides and dimeric peptides exhibiting beta-galactosidase enzyme activity, as well as methods for producing a dairy product using them, and dairy products made by such methods. In some embodiments, the peptides exhibit beta-galactosidase enzyme activity at low and/or high temperatures.

In order to grow on milk, lactose hydrolysis is a good way for lactic acid bacteria to obtain glucose and galactose as carbon source. Lactase (beta-galactosidase; EC 188.8.131.52) is the enzyme that performs the hydrolysis step of the milk sugar lactose into monosaccharides. The commercial use of lactase is to break down lactose in dairy products. Lactose intolerant people have difficulties to digest dairy products with high lactose levels. It is estimated that about 70% of the world's population has a limited ability to digest lactose. Accordingly, there is a growing demand for dairy food products that contain no or only low levels of lactose.
Lactases have been isolated from a large variety of organisms, including microorganisms like Kluyveromyces and Bacillus. Kluyveromyces, especially K. fragilis and K. lactis, and other fungi such as those of the genera Candida, Torula and Torulopsis, are a common source of fungal lactases, whereas B. coagulans and B. circulans are well known sources for bacterial lactases. Several commercial lactase preparations derived from these organisms are available such as Lactozym® (available from Novozymes, Denmark), HA-Lactase (available from Chr. Hansen, Denmark) and Maxilact® (available from DSM, the Netherlands), all from K. lactis. All these lactases are so-called neutral lactases having a pH optimum between pH 6 and pH 8, as well as a temperature optimum around 37° C. When such lactases are used in the production of, e.g. low-lactose yoghurt, the enzyme treatment will either have to be done in a separate step before fermentation or rather high enzyme dosages have to be used because their activity will drop as the pH decreases during fermentation. Also, these lactases are not suitable for hydrolysis of lactose in milk performed at high temperature, which would in some cases be beneficial in order to keep the microbial count low and thus ensure high milk quality. Furthermore, the known lactases would not be suitable for use in a desired process for the production of ultra-heat treated (UHT) milk, wherein enzymes were added prior to the UHT treatment.
WO2010092057 and WO0104276 relates to cold-active beta-galactosidases. WO07110619 relates to beta-galactosidase with high transgalactosylating activity, whereas WO2009071539 relates to beta-galactosidase with lower transgalactosylating activity.

Lactase enzymes with improved properties


An engine 1 is provided with a fuel injection valve 11 for injecting fuel to be supplied to each of cylinders #1 to #4. An electrical control unit (ECU) 30 increases or decreases both a fuel injection amount and an exhaust gas recirculation (EGR) amount so as to quickly approximate torque variation of the engine 1 to a target value. By increasing or decreasing the fuel injection amount so that the target value of the torque variation is in an area of the EGR amount in which the torque variation does not excessively change with the change of the EGR amount, the torque variation is prevented from being largely displaced from the target value on the increase side.

Conventionally, in general car engines, fuel is injected from a fuel injection valve into an intake port, so that a homogeneous mixture of the fuel and the air is supplied to a combustion chamber. In such engines, an intake path is opened or closed by a throttle valve which is activated in response to the operation of an accelerator. The opening or closing of the intake path serves to adjust the amount of intake air (resultantly, the amount of the homogeneous mixture of the fuel and the air) supplied to the combustion chamber of the engine, thus to control the engine power.
In such a so-called homogeneous burning technique, however, a large intake negative pressure is generated due to the throttling operation of the throttle valve, thereby increasing pumping loss and reducing the efficiency. A technique called "stratified burning" which overcomes the above problem is known, where the throttling of a throttle valve is reduced and fuel is directly supplied to a combustion chamber. This enables combustible mixed air to exist in the vicinity of an ignition plug, increasing the air fuel ratio in this region to improve the ignitability.
An engine employing the above technique is provided with a fuel injection valve for stratified burning which injects fuel directing to the region in the vicinity of the ignition plug. During a low load of the engine, fuel is injected from the fuel injection valve for stratified burning so that the fuel is locally supplied in the vicinity of the ignition plug. Simultaneously with this local supply of the fuel, a throttle valve is opened, to realize the "stratified burning" described above. This enables reducing fuel consumption as well as reducing pumping loss.
Such an engine is provided with an exhaust gas recirculation (EGR) mechanism to achieve emission reduction, as in a device described in Japanese Laid-Open Publication No. 7-119513, for example. Such an EGR mechanism includes an EGR path for associating an exhaust path and an intake path of the engine with each other and an EGR valve for opening or closing the EGR path. When the engine operates in a low load range, the opening/closing of the EGR valve is controlled so that as the load increases the amount of exhaust gas to be recirculated (the EGR amount) is reduced. This control enables reducing the air excess rate and thus improving the performance of cleaning of nitrogen oxide in a catalyst device.
The above publication also discloses a technique where torque variation (output variation) of an engine is approximated to a target value by controlling the EGR amount, so as to achieve both suppression of torque variation and reduction of emission. More specifically, when the torque variation exceeds a target value, the EGR amount is decreased to suppress the torque variation to below the target value. When the torque variation becomes less than the target value, the EGR amount is increased to approximate the torque variation to the target value, and thus to reduce the amount of nitrogen oxide emitted from the engine.
During the above "stratified burning", the relationship between the EGR amount and the torque variation is as shown in FIG. 22 when the fuel injection amount is fixed. More specifically, in an area of a smaller EGR amount, the change in torque variation with increase or decrease of the EGR amount is small. In an area of a larger EGR amount, the change in torque variation with increase or decrease of the EGR amount is large. The reason why the change in torque variation with increase or decrease of the EGR amount is small in the area of a smaller EGR amount is that, since a mixed gas with a high-concentration fuel exists in the vicinity of an ignition plug in the "stratified burning", the combustion state of such a mixed gas is not easily changed with a change in the EGR amount.
In the above relationship between the EGR amount and the torque variation, assume that the EGR amount is at point A in FIG. 22. In this case, the torque variation is considerably smaller than the target value. In order to approximate the torque variation to the target value, the control is made to increase the EGR amount. This results in the torque variation shifting as shown by the solid line in FIG. 22 from point A toward the target value, i.e., point B.
In the above case, however, the torque variation increase only at a small change rate as the EGR amount is increased in the area of a smaller EGR amount as described above. It is therefore impossible to quickly approximate the torque variation to the target value.
On the contrary, in the area of a larger EGR amount, the torque variation excessively shifts as the EGR amount changes. This makes it difficult to correctly control the torque variation to be approximated to the target value. To describe more precisely, due to a response delay of the change in the EGR amount, the EGR amount does not immediately change in response to a change in the opening of the EGR valve. Accordingly, when the torque variation exceeds the target value (arrow C in FIG. 22), the torque variation exhibits a sharp increase from the target value due to the response delay at the start of decrease of the EGR amount. Such an excessive torque variation may cause a reduction of drivability.
In recent years, in order to reduce fuel consumption, the air fuel ratio of a homogeneous mixed gas to be supplied to a combustion chamber is made larger than a theoretical air fuel ratio, and swirl is generated in the mixed gas, so as to perform "lean burning".
During the "lean burning", as the EGR amount increases, the ignitability of the mixed gas decreases and simultaneously the flame speed in the mixed gas decreases. Accordingly, if the EGR amount excessively increases, the mixed gas may not be ignited or may fail to be burned out if ignited, resulting in increasing the torque variation of the engine.
During the "lean burning", also, the amount of fuel included in the mixed gas per unit volume decreases. As a result, the torque variation of the engine increases or decreases in response to a change in the fuel injection amount from a fuel injection valve. For example, the torque variation of the engine increases when the fuel injection amount falls below a required amount due to a design size tolerance for the fuel injection valve and the like. On the contrary, the torque variation of the engine decreases when the fuel injection amount exceeds the required amount due to a design size tolerance for the fuel injection valve and the like.
According to the above conventional technique, when the torque variation increases due to a decrease of the fuel injection amount below the required amount due to a design size tolerance and the like, it is attempted to suppress the torque variation by decreasing the EGR amount to be approximated to the target value. However, since the above increase of the torque variation is caused by the decrease of the fuel injection amount due to a design size tolerance for the fuel injection valve and the like, such torque variation is not suppressed even if the EGR amount is decreased. Surging generated in the engine due to the torque variation is not suppressed, either. Moreover, since the emission amount of nitrogen oxide (NO.sub.x) increases due to the decrease of the EGR amount, emission is worsened.

Combustion control device for internal combustion engine


Attenuating element consisting of a head part, neck part and base part. The neck part is narrower compared to the head part and neck parts of adjacent attenuating elements form a continuous channel. Via the head parts, this channel is connected so that the cross-sectional area of the head parts is smaller than the cross-sectional area of the base parts. In this way, a water-control structure, sound-attenuating wall and the like can be provided. Such attenuating elements can be produced in a very simple manner by dividing them vertically and producing each of the vertical parts from concrete in a mould.

1. Field of the Invention
The invention relates generally to an attenuating element.
2. Description of the Related Art
An attenuating element of this type can be used to attenuate the action of the waves, for example on water-control structures, such as dykes. In addition, the attenuating element according to the present invention can be used for attenuating sound waves, wherein a number of attenuating elements form a sound-attenuating barrier or wall. Other possibilities to attenuate the energy of waves can be achieved by means of attenuating elements, for example the removal of large quantities of water.
In the prior art, series of blocks which have been placed against one another have been used as attenuating elements which are used for dykes, wherein the upper surfaces of such blocks have been placed in a more or less adjoining manner. It has been found that it is possible to achieve an improvement if apertures are provided on the upper side into which the water can be moved. As a result thereof, the flow of the water flowing onto the dyke is disrupted and the attenuating action is increased.
However, in the prior art this requires highly complex elements which are associated with correspondingly high production costs.
US 2002/0025231 discloses an attenuating element assembly consisting of a series of attenuating elements with a common base part. Such a structure is very difficult to install, can certainly not be handled by hand and does not provide a possibility for modifications to the shape of the body located underneath, such as a dyke body or another wall. Due to the common base part, there is no interaction with the remainder of the dyke.
U.S. Pat. No. 5,556,230 discloses a coastal defence system consisting of a number of elements with narrowed and enlarged parts which are placed against one another in turns. Although this does make it possible to follow the shape of a dyke body, it only achieves limited additional attenuation, if any. Such a structure is only suitable for preventing erosion of material.
All structures of this kind have the drawback that they are so heavy that they cannot be readily installed by hand and, in addition, are very difficult to produce from concrete material. More particularly, they require a very complicated mould, as a result of which it is not possible to produce them at high speed and on a large scale at relatively low cost.
WO 03/076727 discloses an ecological block which can be used on river banks and is provided with cavities which are to be filled with vegetation. The block is substantially rectangular and, in a first embodiment, is substantially flat on two opposite sides. In another variant (FIG. 6), one side thereof is flat and the opposite side is provided with a head part and a base part. It is intended to place the flat side against a wall.

This patent is subject to a terminal disclaimer.


Disclosed is a gate mechanism (100) mounted on a side frame (210) of a financial automated machine cash slot (230a), wherein the gate mechanism comprises a gate (10), a driving mechanism (20) and a pair of locking mechanisms (30) controlling the gate (10), with the lower end of the gate (10) being connected with a front end (240a) of the frame side (240) in such a way as to be able to slide up and down, and the upper end of the gate (10) being connected with an upper end (240b) of the frame side (240) in such a way as to be able to slide back and forth, and the driving mechanism (20) driving the upper end of the gate (10) in sliding back and forth at the upper end (240b) of the frame side (240), while causing the lower end of the gate (10) to move up and down at the front end (240a) of the frame side (240), thus achieving opening and closing of the gate (10). The gate mechanism (100) can reduce the motion enveloping space of the gate (10) and has the advantage of a compact structure.

The financial self-service device has been widely used in everyday life, and can provide 24-hour continuous service and bring significant convenience for customers. However, for the unattended automated teller machine, there exists a risk that the money of is the depositor may be stolen or intercepted by criminals. Therefore, the application security of the financial self-service device is causing more and more attention. For providing a violence-preventing protection for the financial self-service device, there are several structural embodiments which can realize the function of a shutter of the financial self-service device, such as opening, closing and locking function.
The shutter structure of some commercially available financial self-service devices mainly has two action principles, that is, a sliding manner, in which the shutter is placed horizontally or tilted at a certain degree, and a pivoting manner. One solution uses a principle of the shutter mechanism sliding back and forth in a certain plane. For example, as disclosed in a patent application No. 200910041871.7 titled "A safe shutter device and an automated teller machine having the same" with the publication No. CN101684713A, two planar sliding grooves are respectively provided at two sides of the shutter of the safe shutter device, the shutter mechanism is driven by a force to slide back and forth along a fixed sliding groove, and two extreme positions of the planar sliding groove respectively correspond to an open state and a close state of the shutter mechanism. The major problem of such shutter mechanism is that, because the mounting space is relatively large due to the alternative open and close states, the shutter mechanism is generally adapted to be placed horizontally or tilted.
In another solution, the shutter mechanism rotates around an axis to realize the open and close actions. For example, in the automatic shutter device disclosed in the patent application No. 200910193709.7 with the publication No. CN101840596A, one end of the shutter of the device is fixed on a rotating axis, and the shutter is driven by a force to rotate around the axis by a certain angle from an initial angle, such that the shutter is in close and open states respectively. The major problem of such shutter mechanism is that, the enveloping space of the spatial motion is relatively large due to the motion manner, thus enough space is required for realizing the action of the shutter, and if there is a requirement on the space limitation, the application of such shutter mechanism is not convenient. A shutter mechanism on the product LOBBY90 of Wincor Company, which also rotates around an axis, solves the problem of the large motion enveloping space, however the arc shutter plate causes a lot of inconveniences during the overall mounting and fitting process.
Further, for some through-the-all or outdoor placed financial self-service devices, the transaction interface is generally exposed, which requires the shutter having a certain rainproof function.
Therefore, it is urgent to provide a shutter mechanism which is able to reduce the enveloping space of the motion of the shutter and has a compact structure.


A mechanical rotary coupler for first and second coral shafts having slotted ends proximate one another, the coupler being formed of an elongated blade of resilient material longitudinally slit and folded on itself transverse to the slit to provide first and second resilient V-shaped strips connected by bridging portions at the ends of the blade, the strips respectively being adapted to engage the slots in the ends of the first and second shafts at regions on the strips spaced from the bridging portions.

The invention disclosed herein relates generally to mechanical couplers for rotatable shafts, and more particularly to such couplers of a design which can be simply formed of inexpensive sheet material in a configuration which tolerates shaft misalignment and permits limited overtravel of one shaft relative to another.
Coupler devices of wide variety of designs for generally aligned rotatable shafts are well known. These designs range from simple solid couplers which rigidly connect one shaft to another end to end, to couplers employing a variety of resilient elements for permitting limited relative motion and/or misalignment between the coupled shafts.
One common resilient coupler design type employs a pair of flanges or hubs rigidly fixed to adjacent ends of the shafts, the flanges or hubs being connected to one another through elastomeric material, springs or a functional equivalent. In such an arrangement, the coupler is normally formed of a plurality of parts, some of which may be most advantageously formed by machining or casting. The number and form of the component parts results in a coupler which is relatively complex and expensive.
It is known to simplify this arrangement by appropriately configuring the spring and ends of the shafts so as to eliminate the flanges or hubs. A particular implementation may effectively involve a cylindrical coil spring aligned with the axis of the shafts and connected at opposite ends thereof to the separate shafts. However, the obvious ways of connecting the spring to the shafts in such an implementation either require special forming which may be disadvantageous for production, or are difficult to assemble and disassemble, or do not provide reliable attachment which is adequately free of backlash, or have some other associated disadvantage.
Although existing rotary coupler designs are satisfactory for many applications, a continuing need exists for simpler, less expensive couplers which are adaptable to a variety of applications, and exhibit characteristics such as freedom from backlash, tolerance of shaft misalignment and overtravel of one shaft relative to the other, accommodation of shafts of unequal sizes, simple single piece construction capable of inexpensive and easy manufacture, etc.

Rotary coupler with shaft misalignment and overtravel tolerance


Various embodiments of irrigation fluid collection basins are disclosed. The basin is configured for irrigation of a wound or fracture generally located on a portion of the human anatomy, such as a knee, shin, calf, elbow, or otherwise. The basin can include two recesses located on opposite sides of the basin along an upper periphery of the basin. The recesses are configured to receive the portion of the human anatomy (e.g., a leg) at two regions proximal to the location of the wound or fracture, so that the wound or fracture is located over the cavity of the basin. The recesses can be sized so that one is broader than the other, the broader recess configured to comfortably accommodate a larger region of the leg and the other recess configured to comfortably accommodate a smaller region of the leg.

1. Field of the Invention
The present inventions are directed to surgical equipment, and in particular, equipment used for irrigating wounds on the human body.
2. Description of the Related Art
Basins are widely used in the medical field to collect irrigation fluid during an irrigation procedure of a wound on a patient. The wound may consist of a laceration or a cut that breaks a skin surface. Fractures may also require irrigation procedures. Medical personnel irrigate such wounds to flush out any contaminants from the wound prior and/or subsequent to conducting a medical procedure on the wound area.
Irrigation procedures are common in the medical field, particularly in the field of orthopedic surgery, due to the fact that wounds are fairly common injuries, many of them requiring treatment at a medical facility, particularly deep cuts. Therefore, medical facilities tend to maintain a significant number irrigation kits ready for use during surgical procedures. This is also true for emergency room facilities, where patients with wounds requiring immediate treatment generally arrive, and where the number of patients that will arrive throughout a day is impossible to predict. However, the use of irrigation kits is not limited to emergency room procedures and includes other medical procedures as well.
Existing irrigation kits typically include at least one basin to collect the irrigation fluid wrapped in plastic or other material preventing dirt or other contamination from coming in contact with the basin. Wrapping the kit in plastic also allows the kits to be kept separate from each other, even if the kits are stacked one on top of another. Moreover, each kit generally includes various components necessary for an irrigation procedure so that medical personnel usually need not open more than one irrigation kit to perform an irrigation procedure. The components, such as the basins, are usually sterilized.
When a wound requires irrigation, medical personnel open an irrigation kit to take the basin out of the kit, and place the basin in an area below the region of the patient's body where the wound is located. Then, with an irrigation device, the medical personnel directs an irrigation fluid, such as, but without limitation, water, saline, or a solution including antibacterials and/or antibiotics at the wound area to remove contaminants, tissue and/or bone fragments from the wound.
The irrigation device preferably delivers irrigation fluid at a pressure and flow rate sufficient to effectively clean the wound area. The irrigation device may have a shield connected to the device to reduce the amount of irrigation fluid that splashes off the wound field and/or in the direction of the treating medical personnel or on the floor. The shield can further be used to focus the direction of the irrigation fluid on the wound when the device is placed close to the wound area.
Throughout the irrigation procedure, medical personnel adjust the location of the basin to collect the irrigation fluid after it strikes the wound area. For example, one may place the basin just below the body part where the wound is located. Alternatively, one may place the basin in a location to which the irrigation fluid flows after striking the wound area.

Irrigation fluid collection basins that receive portions of the human anatomy


An improved solution for Web services is provided. In an embodiment of the invention, a method for providing a virtual Web service includes: providing a Web service gatekeeper, where the gatekeeper acts as an access point to multiple private internal enterprise environments; and then the gatekeeper receives a request for access to one, or more, of these private internal enterprise environments.

Typically, Web and other local services are made freely available, or are accessible on a per-fee basis uniquely tailored to the particular system and service in question. For example, when a corporation wishes to provide a Web service for aa fee, they must create unique infrastructure to both secure (from unpaid usage) and advertise their Web service. Another example is when a business traveler passes through an airport and wishes to access a wireless network. Every time the payment model and/or terms of the Web service are unique to the particular situation, it causes frustration to the user, as well as provides no mode for flexible pricing (e.g., an all-day pass for airports across a certain geography, etc.).
The aforementioned inefficiencies also may lead to higher service pricing because more development time is needed to deploy a particular service with its associated fee. This also leads to fragmentation because a heightened user intervention is required to gain access to the multitude of available services. As a result, no software can be made to utilize multiple Web services, thereby defeating an aspect of for-fee service oriented architecture.
In view of the foregoing, there is an opportunity to improve upon the related art.

International Business Machines Corporation


There is provided composition comprising i) a thermoplastic polymer ii) a compound having the formula

The manufacturing properties of thermoplastic polymers, for example the extruding properties of such polymers, is often modified/enhanced by the addition of plasticisers thereto. As acknowledged in the prior art, such as in U.S. Pat. No. 4,426,477, there is a tendency toward avoiding the commonly used plasticisers such as dioctyl adipate (DOA) and phthalate plasticisers such as dioctyl phthalate (DOP). The safety of these plasticisers has been called into question, particularly in certain applications.
U.S. Pat. No. 4,426,477 discloses plasticisers based on glycerol esters. The plasticisers consist of compounds prepared by the acylation of glycerol. The compounds comprises triesters, wherein approximately two of the acyls have two carbons and the remaining one acyl has from 10 to 14 carbons. The compounds of U.S. Pat. No. 4,426,477 provide a plasticising effect. However, in certain applications the plasticisers have a volatility such that they may migrate out of the thermoplastic polymer in which they are incorporated, such as PVC.
In a first aspect the present invention provides a composition comprising i) a thermoplastic polymer ii) a compound having the formula
wherein R1, R2 and R3 are independently selected from an acyl group or a hydrogen atom, wherein at least one of R1, R2 and R3 is an acyl group (a short acyl group) having from 2 to 6 carbon atoms, wherein at least one of R1, R2 and R3 is a branched chain acyl group (a long acyl group) consisting of or having a saturated chain having 10 to 20 carbon atoms and a hydrophilic branch group.
Preferably two of R1, R2 and R3 are short acyl groups as described above and the other of R1, R2 and R3 is a long acyl group as described above. In this aspect, the compound may be of the formula
Preferably the hydrophilic branch group is a group selected from acyl and derivatives thereof. Preferred derivatives include groups of the formula xe2x80x94O-acyl.
Preferably the hydrophilic branch group is a group of the formula
wherein p is from 0 to 4 or 0 to 3.
In a preferred aspect of the present invention the chain of the long acyl group consists of or has a saturated chain having 14 to 20 carbon atoms. In a more preferred aspect, the chain of the long acyl group consists of or has a saturated chain having 16 to 20 carbon atoms
Preferably the long acyl group is of the formula
wherein n is from 10 to 20 and m is 2n, and wherein p is from 0 to 4 or 0 to 3.
Preferably n is from 16 to 20, more preferably from 16 to 18, yet more preferably 17.
Preferably the group CnHm is a straight chain hydrocarbon group.
In a highly preferred aspect the long acyl group is a group of the formula
wherein x is from 7 to 10, preferably x is 10, and y is 2x, and wherein p is from 0 to 4 or 0 to 3, preferably p is 0.
Preferably the group CxHy is or comprises a straight chain hydrocarbon group.
In a highly preferred aspect the long acyl group is a group of the formula
In a preferred aspect of the present invention the short acyl group is or comprises an acyl group having from 2 to 5 carbon atoms. In a more preferred aspect, the short acyl group is or comprises an acyl group having 2 carbon atoms. The short acyl group is preferably of the formula
Preferably the short acyl group and the hydrophilic branch group contain the same number of carbon atoms. In a highly preferred aspect the hydrophilic branch group is a group of the formula
and the short acyl group is of the formula
wherein p=q and is from 0 to 4 or 0 to 3.
In certain aspects, it is desirable for the short acyl groups to be present in a maximum amount with respect to the total amount glycerol and esters thereof present in the composition. Preferably the short acyl group is present in an amount on average, of no greater than 2 moles per mole of glycerol and esters thereof present in the composition.
In certain aspects, it is desirable for the long acyl groups to be present in a minimum amount with respect to the total amount glycerol and esters thereof present in the composition. Preferably the long acyl group is present in an amount, on average, of at least 0.4 moles, preferably from 0.9 to 2 moles, more preferably from 0.9 to 1 moles per mole of glycerol and esters thereof present in the composition.
It may also be preferred for the majority of the glycerol present in the composition to be fully acylated Accordingly, in a preferred aspect the total amount of acyl groups is, on average, 2.7 to 3.0 moles per mole of glycerol and esters thereof
The compound of the present invention may be prepared by interesterification between glycerol and one or more oils, including natural oils and hardened natural oils followed by acylation. Thus, the compound of the present invention may be the product of a two part process comprising (i) an interesterification between glycerol and an oil selected from castor oil, including hardened castor oil, unhardened castor oil and mixtures thereof, and (ii) acylation.
Thus in a further aspect the present invention provides a process for the preparation of a compound having the formula
wherein R1, R2 and R3 are independently selected from an acyl group or a hydrogen atom; wherein at least one of R1, R2 and R3 is an acyl group (a short acyl group) having from 2 to 6 carbon atoms or from 2 to 5 carbons; wherein at least one of R1, R2 and R3 is a branched chain acyl group (a long acyl group) consisting of or having a chain having 10 to 20 carbon atoms and a hydrophilic branch group; the process comprising the steps of:
(i) interesterification between glycerol and triglyceride compound having the formula
wherein each of R1, R2 and R3 is a fatty acid group consisting of or having a chain having 10 to 20 carbon atoms, to provide a composition comprising glycerol, monoglyceride, diglyceride and/or triglyceride;
(ii) optionally isolating the monoglyceride and/or diglyceride from the composition;
(iii) acylating the monoglyceride and/or diglyceride or the composition containing the same.
In the process of the present invention the chain having 10 to 20 carbon may be saturated or unsaturated.
The process of the present invention may utilise, for example, castor oil or hardened castor oil. The compound of the present invention may be prepared from hardened castor oil. A typical fatty acid profile of castor oil and hardened castor oil is given below.
The nomenclature in parenthesis is Cxx:y where xx is the fatty acid carbon number and y indicates number of double bonds. Ricinoleic acid, hard (also known as 12-hydroxy stearic acid) has a hydroxyl group (OH) on the 12th carbon.
In this aspect the product based on the castor oil, or indeed a product based on another oil, may be synthesised as follows. These synthetic routes are given by way of example only. Other routes would be appreciated by a person skilled in the art
Route 1
Castor oil and glycerol are reacted to produce an equilibrium mixture of different glycerol esters. This mixture can be distilled and subsequently acetylated to provide the desired end products. This route is illustrated below.
The second, step comprising the removal of the glycerol followed by distillation is optional. In other word the mixture of glycerol, glycerol monoester, glycerol diester, and glycerol triester may directly acetylated with acetic anhydride to form a mixture of acetylated products. The acetylated monoglyceride or the mixture of acetylated products may have a yellowish color. The yellowish product may be distilled. The distilled product is clear.
Depending on the ratio between castor oil and glycerol and depending on the reaction conditions, the described process provides different glycerol ester mixtures. The glycerol monoester is typically 45-65% and may be up to 65%. This is achieved in a typical reaction mixture. Such a composition may then, optionally, be fir processed to provide a product having a higher purity, for example a monoester content of above 90%.
Typical further processing would involve the removal of glycerol and the subsequent distillation of the resulting mixture. By means of this further processing an end product with a typical glycerol monoester content above 90% may be provided. The fatty acid profile of the end product would reflect the fatty acid profile of the castor oil stating material. This process, namely the production of a glycerol monoester, is described in Bailey""s Industrial Oil and Fat Products, vol. 4, Fifth ed. P.572 ff
The glycerol monoester may then be reacted with an excess of acetic anhydride resulting in acetic acid being esterified with the glycerol ester. Any excess of acetic anhydride is removed from the reaction mixture. The end product is a fully acetylated monoglyceride based on castor oil (which may be hardened or unhardened castor oil).
Route 2
Route 2 provides a further manner in which acetylated monoglycerides based on, for example, hardened and unhardened castor oil may be synthesised. An acetylated monoglyceride may be produced by the reaction of castor oil with acetic anhydride to provide an acetylated castor oil, The acetylated castor oil is then reacted with triacetin to produce acetylated monoglycerides based on hardened and unhardened castor oil. This process is illustrated below.
Castor oil is reacted with acetic anhydride to produce an acetylated castor oil. The degree of acetylation may be defined as castor oil containing at least one acetic acid group and up to the average of three acetic acid groups per mole of castor oil. The acetylated castor oil is then reacted with triacetin to produce a reaction mixture of mainly acetylated glycerolesters with a fatty acid profile similar to that of the castor oil starting material. This mixture is purified by distillation. The end product is an acetylated monoglyceride based on hardened or unhardened castor oil.
A process for obtaining acetylated glycerol esters from the reaction of a triglyceride and triacetin is described in the Danish Patent No. 93 786. The synthesis of acetylated monoglycerides based on castor oil from the starting materials acetylated castor oil and triacetin is analogous to that process.
In a highly preferred aspect the compound of the present invention is selected from compounds of the formula
These preferred compounds may be prepared and are preferably prepared in accordance with Route 1 or Route 2 above.
The above compounds are specific compounds which may be provided by the present invention. In a broad aspect, the present invention further provides a compound of the formula
wherein two of R4, R5, and R6 are of the formula
wherein for each of the two of R4, R5, and R6 q is independently selected from 0 to 4 or from 0 to 3; and the other of R4, R5, and R6 is a branched group of the formula
wherein n is from 10 to 20 and m is 2 n, and wherein p is from 0 to 4 or from 0 to 3.
Preferably q is 0. More preferably for both of the two of R4, R5, and R6 q is 0.
Preferably n is from 16 to 20, more preferably from 16 to 18, yet more preferably 17.
Preferably the group CnHm is a straight chain hydrocarbon group.
In a preferred aspect the branched group is a group of the formula
wherein x is from 7 to 10, preferably x is 10, and y is 2x, and wherein p is from 0 to 4 or from 0 to 3, preferably p is 0.
Preferably the group CxHy is a straight chain hydrocarbon group.
The compounds provided by the present invention may be incorporated in a composition comprising a thermoplastic polymer. Accordingly in a further aspect the present invention provides a composition comprising a compound as defined above and a thermoplastic polymer.
The thermoplastic polymer of the compositions of the present invention may be or comprise a vinyl chloride polymer or a vinyl chloride copolymer selected from vinyl chloride/vinyl acetate copolymer, vinyl chloride/vinylidene chloride copolymer, vinyl chloride/ethylene copolymer and a copolymer prepared by grafting vinyl chloride onto ethylene/vinyl acetate copolymer, or a mixture thereof.
In a preferred aspect the thermoplastic polymer is or comprises a polymer blend of a thermoplastic polymer, preferably a thermoplastic polymer as defined above, and a second polymer. Preferably, the second polymer is a methacryl polymer or an acrylonitrile-butadiene-styrene polymer.
The compositions of the present invention may be formulated in any manner to provide the required plasticising properties of the compound. In a particular aspect the composition of the present invention comprises the compound in an amount of 1 to 100 parts by weight per 100 parts by weight of the thermoplastic polymer.
As disclosed above the present invention provides a process for the preparation of a compound having the formula
wherein R1, R2 and R3 are independently selected from an acyl group or a hydrogen atom, wherein at least one of R1, R2 and R3 is an acyl group (a short acyl group) having from 2 to 5 or from 2 to 6 carbon atoms, wherein at least one of R1, R2 and R3 is a branched chain acyl group (a long acyl group) consisting of or having a chain having 10 to 20 carbon atoms and a hydrophilic branch group, the process comprising the steps of: (i) interesterification between glycerol and triglyceride compound having the formula
wherein each of R1, R2 and R3 is a fatty acid group consisting of or having a chain having 10 to 20 carbon atoms, to provide a composition comprising glycerol, monoglyceride, diglyceride and/or triglyceride; (ii) optionally isolating the monoglyceride and/or diglyceride from the composition; (iii) acylating the monoglyceride and/or diglyceride or the composition containing the same.
In the process of the present invention the chain having 10 to 20 carbon atoms may be saturated or unsaturated.
In preferred aspects the compound prepared by the process is a compound of the present invention as described herein. In a broader aspects
the long acyl group is of the formula
wherein n is from 10 to 20 and m is selected from 2n, 2nxe2x88x922, 2nxe2x88x924 and 2nxe2x88x926, and wherein p is from 0 to 4 or from 0 to 3.
Preferably m is 2n or 2nxe2x88x922.
Preferably the group CnHm is a straight chain hydrocarbon group. The straight chain hydrocarbon group may be saturated or unsaturated. The straight chain hydrocarbon group may contain a single xe2x80x94Cxe2x95x90Cxe2x80x94 bond.
the long acyl group is a group of the formula
wherein x is from 7 to 10, preferably x is 10, and y is 2xxe2x88x922, and wherein p is from 0 to 4 or from 0 to 3, preferably p is 0.
Preferably the group CxHy is a straight chain hydrocarbon group. The straight chain hydrocarbon group may be saturated or unsaturated. The straight chain hydrocarbon group may contain a single xe2x80x94Cxe2x95x90Cxe2x80x94 bond.
the long acyl group is a group of the formula
the compound is of the formula
the compound is of the formula
wherein two of R4, R5, and R6 are of the formula
wherein for each of the two of R4, R5, and R6 q is independently selected from 0 to 4 or from 0 to 3
and the other of R4, R5, and R6 is a branched group of the formula
wherein n is from 10 to 20 and m is selected from 2n, 2nxe2x88x922, 2nxe2x88x924 and 2nxe2x88x926, and wherein p is from 0 to 4 or from 0 to 3. Preferably q is 0. More preferably for both of the two of R4, R5, and R6 q is 0. Preferably n is from 16 to 20, more preferably from 16 to 18, yet more preferably 17. Preferably m is 2nxe2x88x922.
the branched group is a group of the formula
wherein x is from 7 to 10, preferably x is 10, and y is 2xxe2x80x942, and wherein p is from 0 to 4 or from 0 to 3, preferably p is 0.

Abstract: wherein R1, R2 and R3 are independently selected from an acyl group or a hydrogen atom, wherein at least one of R1, R2 and R3 is an acyl group (a short acyl group) having from 2 to 6 carbon atoms, wherein at least one of R1, R2 and R3 is a branched chain acyl group (a long acyl soup) consisting of a saturated chain having 10 to 20 carbon atoms and a hydrophilic branch group.


An information processing apparatus including a display unit that is capable of displaying an operation screen provided by a server notifies the server of a content of an instruction from a user via an operation screen provided by the server and displayed on the display unit, and if the server requests execution of a process based on the notified content of the instruction, executes the requested process. The information processing apparatus determines an operation screen displayed on the display unit after notifying the server, and if the operation screen provided by the server is displayed on the display unit, performs control to execute the process requested by the server, and if another operation screen is displayed on the display unit instead of the operation screen provided by the server, performs control to not execute the process requested by the server.

1. Field of the Invention
The present invention relates to an information processing apparatus that includes a display unit capable of displaying an operation screen provided by a server.
2. Description of the Related Art
An information processing apparatus such as a personal computer (PC) which is connected to a server on a network and displays on a display unit therein an operation screen provided by the server is conventionally known. For example, there is a system in which a web server included in the information processing apparatus displays the operation screen acquired from a web server on the network.
In such a case, the web browser in the information processing apparatus requests the web server to provide the operation screen. A web application in the web server then responds to the request received from the information processing apparatus by transmitting a hypertext markup language (HTML) file for the web browser to display the operation screen. The web browser in the information processing apparatus analyzes the received HTML file and displays the operation screen based on the description of the HTML file.
Further, if a user inputs an instruction via the operation screen displayed on the web browser, the web browser notifies the web server of the input instruction. The web application on the web server that receives the notification then executes a process according to the input instruction.
Recent multifunction peripherals (MFP) which include a scanner and a printer also include the above described web browser. The MFP thus displays on the web browser the operation screen provided by the web server based on the above described procedure and receives various instructions from the user.
Further, Japanese Patent Application Laid-Open No. 2006-127503 discusses a technique in which the web server provides the operation screen to which the user inputs instructions for using various functions included in the MFP. In other words, a user of the MFP inputs an instruction to the MFP via the operation screen displayed on the web browser. The web browser of the MFP then notifies the web server of the input instruction.
Upon receiving such a notification, the web server requests the MFP to perform various processes according to the content of the instruction input by the user. The MFP which receives such a request thus performs the requested process. As a result, it becomes unnecessary to store in the MFP all pieces of operation screen information for operating the MFP. Further, the content of the operation screen can be easily changed on the web server.
The technique discussed in Japanese Patent Application Laid-Open No. 2006-127503 once notifies the server of the instruction input via the operation screen displayed on the display unit in the information processing apparatus. The information processing apparatus then performs various processes according to the request that is returned from the server. However, in such a case, problems are generated as described below.
More specifically, the user may operate on the information processing apparatus and switch the operation screen displayed on the display unit to another operation screen after the information processing apparatus notifies the server of the instruction input via the operation screen and is then requested by the server to execute the process. For example, after the information processing apparatus notifies the web server of the instruction input via the operation screen displayed on the web browser, the user may close the web browser and then switch to another operation screen provided by the application in the information processing apparatus. Further, the user may newly display an operation screen provided by the application in the information processing apparatus in front of the web browser while the web browser continues to be activated.
In the above described cases, if the process requested by the server requires a user operation to be performed after the process is executed, the user operation is not performed even when the process is executed. Execution of the process may thus become useless. For example, in one case it is assumed that a process requested by the server is to read a document, and it is necessary for the user to preview and confirm image data acquired by executing the process. In such a case, the user cannot preview and confirm the image data if the web browser is closed. As a result, the processes of previewing and confirming the image data and thereafter are not performed, so that the reading process becomes useless.
Further, for example, it is assumed that when a process requested by the server is executed, the information processing apparatus displays a screen indicating that the "process is being executed" based on screen information stored therein. In such a case, when the user is operating on the new operation screen, the screen may be switched (or displayed by overwriting on the frontmost screen) to the screen indicating that the "process is being executed" simultaneously as the process which is requested in delay from the server is started. The user operation thus becomes interrupted.

Information processing apparatus, control method of information processing apparatus, and storage medium