A voltage adjustment circuit of a semiconductor memory apparatus includes a control voltage generating unit configured to distribute an external voltage for selectively outputting a plurality of distribution voltages as a control voltage in response to a control signal, the plurality of the distribution voltages each have different voltage levels, a comparing unit configured to include a voltage supply unit configured to control an external voltage supplied to a first node and a second node if a level of an output voltage is higher than a level of a reference voltage in response to a level of the control voltage, and a detection signal generating unit configured to drop potential levels of the first and second nodes according to the levels of the output voltage and the reference voltage, and to output the potential level of the second node as a detection signal, and a voltage generating unit configured to drive the external voltage according to a potential level of the detection signal and to output the external voltage as the output voltage.

1. Technical Field
The embodiments described herein relate to a semiconductor memory apparatus, and more particularly, to a voltage level comparison circuit, a voltage adjustment circuit using the voltage level comparison circuit, and a semiconductor memory apparatus using the same.
2. Related Art
In general, a semiconductor memory apparatus drops/reduces an external voltage and generates a voltage having a desired level. Here, a circuit that drops the external voltage and generates a stable voltage is commonly called a voltage regulator or a voltage adjustment circuit. Here, the voltage adjustment circuit has been widely used as an internal voltage generation circuit that generates an internal voltage.
FIG. 1 is a schematic diagram of a conventional voltage adjustment circuit of a semiconductor memory apparatus. In FIG. 1, a voltage adjustment circuit 1 includes a comparison unit 10 and a voltage generating unit 20.
The comparison unit 10 compares levels of a reference voltage Vref and an output voltage V_out, and generates a detection signal 'det'. The comparison unit 10 includes first to fourth transistors P1, P2, N1, and N2. The first transistor P1 is supplied with an external voltage VDD through a source terminal thereof, and supplies a voltage to a first node (node A) that is connected to a drain terminal thereof. The second transistor P2 is supplied with the external voltage VDD through a source terminal thereof, and supplies a voltage to a second node (node B) that is connected to a drain terminal thereof. In the third transistor N1, the output voltage V_out is supplied to a gate terminal thereof, a ground terminal VSS is connected to a source terminal thereof, and the first node (node A) is connected to a drain terminal thereof. In the fourth transistor N2, the reference voltage Vref is supplied to a gate terminal thereof, the ground terminal VSS is connected to a source terminal thereof, and the second node (node A) is connected to a drain terminal thereof.
Here, the detection signal 'det' is output from the second node (node A). In addition, the voltage generating unit 20 drives an external voltage VDD according to a potential level of the detection signal 'det' and generates the output voltage V_out. The voltage generating unit 20 includes a fifth transistor P3, a resistor element R1, and a capacitor C1. In the fifth transistor P3, the detection signal 'det' is supplied to a gate terminal thereof, the external voltage VDD is supplied to a source terminal thereof, and the output voltage V_out is output from a drain terminal thereof. The resistor element R1 and the capacitor C1 are connected in parallel and are connected between the drain terminal of the fifth transistor P3 and the ground terminal VSS. Here, the resistor element R1 and the capacitor C1 that are connected in parallel function as a filter that directs noise of the output voltage V_out to the ground terminal VSS.
In the voltage adjustment circuit 1, if the noise is generated in the external voltage VDD, then the noise is also generated in the output voltage V_out. Specifically, a potential level of the second node (node B), i.e., a potential level of the detection signal 'det', is generated by supplying the external voltage VDD to the second node (node B) when the second transistor P2 is turned ON. Thus, the noise of the external voltage VDD is output as the detection signal 'det' without being removed. Here, the detection signal 'det' that includes the noise is supplied to the gate terminal of the fifth transistor P3 but does not fix a turned-ON degree of the fifth transistor P3. Accordingly, the output voltage V_out that is output from the fifth transistor P3 also includes the noise.

Voltage level comparison circuit of semiconductor memory apparatus, voltage adjustment circuit using voltage level comparison circuit, and semiconductor memory apparatus using the same


A packaging and application device for a product, specifically a cosmetic product. The device includes a receptacle for cleaning the product having a longitudinal axis and a passageway. The receptacle also includes a porous or fibrous applicator element capable of communicating with the product in the receptacle through the passageway. A dispensing element adapted for opening and closing the passageway. The device further contains a dispensing element adapted for opening and closing the passageway and includes a mobile part rotatable about the longitudinal axis between a first position in which the passage is closed and a second position in which the passage is open. The device also includes a closure cap capable of engaging a dispensing element.

The invention disclosed relates to a packaging and application device for a product, in particular a fluid product such as for example nail varnish remover.
Nail vanish remover products are for the most part packaged in bottles and require the use of a cotton wool swab for their application. Wipes impregnated with remover also exist which have the advantage of being more readily transportable. However, these two types of application have the major drawback that it is difficult to remove the varnish from a nail on one hand without damaging the varnish on the other nails of both hands, particularly the hand in which the cotton wool swab or wipe is being held. It often happens that the varnish on a single nail is damaged and it is needed to remove the varnish from this nail only without re-applying varnish to both hands. Similarly, when it is desired to remove the varnish from the toenails, the use of a cotton wool swab or wipe impregnated with remover removes the varnish from the nails of the hand holding the swab or wipe.
For this reason, the need has arisen for an applicator device for a nail varnish remover product which can be used to remove the varnish from a single nail if desired, without removing the varnish from the other nails.
In particular, the need has arisen for a device of the type which includes a receptacle containing the product and an applicator element capable of communicating with the product contained in the receptacle through a passage which can be opened or closed in response to the actuation of an intermediate element placed between the applicator and the receptacle.
Numerous application devices of this type have already been described for other types of products.
In particular, U.S. Pat. No. 4,279,527 describes a device including a bottle onto which is screwed a collar integral with an applicator brush. When the collar is screwed to its maximum extent onto the bottle, the brush is isolated from the inside of the bottle by means of a centre-post which blanks off a communicating aperture between the bottle and brush. To use the device, the user unscrews the collar which moves the aperture away from the centre-post thereby providing sufficient clearance to allow the product to pass from the bottle to the brush. In this device, no closure cap is provided to cover the brush in the closed position so that the brush is not protected.
U.S. Pat. No. 2,409,933 describes in one of the embodiments a bottle including an outlet aperture formed outside the longitudinal axis of the bottle, and which is surmounted by an applicator having an inlet aperture also formed outside the longitudinal axis of the bottle. A cap is provided to cover the applicator in the closed position. In the closed position, the two apertures are offset in such a way that the product cannot reach the applicator. To place the applicator in communication with the product, the user removes the cap then turns the applicator so as to bring the two apertures into alignment with each other. To close the device, the user must first turn the applicator to place the two apertures in the offset position, then replace the cap. Two separate movements are thus necessary both to open and close the device.
In certain devices, the cap serving to protect the applicator is also used to close the passage allowing communication between the inside of the receptacle and the applicator. This is notably the case in the device described in U.S. Pat. No. 3,106,742 wherein a screwing action applied to the closure cap causes the collar integral with the applicator to move downward. However, placement of the applicator in communication with the inside of the receptacle must be done by hand, i.e. the user must first unscrew the cap then pull on the collar in order to open up the passage between the inside of the receptacle and the applicator.
In other devices, opening of the cap causes the applicator to be placed in communication with the product contained in the receptacle. This is notably the case in the devices described in U.S. Pat. No. 5,230,579 and U.S. Pat. No. 3,281,887. In U.S. Pat. No. 5,230,579, the cap is opened by means of a linear movement on the longitudinal axis of the device. When it is opened, the cap causes the movement by friction of an intermediate part carrying the applicator and the clearance of a passage aperture allowing the applicator to communicate with the product contained inside the receptacle. To shut off communication between the applicator and the product inside the receptacle, the user can use the cap to cause the intermediate part carrying the applicator to move downward on the longitudinal axis. In U.S. Pat. No. 3,281,887, the cap is screwed onto the receptacle, under a bellows formed in the neck of the receptacle. In this position, the bellows is compressed. When the cap is unscrewed, the bellows expands and a passage aperture is cleared enabling the applicator to communicate with the product inside the receptacle. When the cap is screwed back onto the receptacle, the bellows is compressed and the aperture recloses.
In these two devices, the applicator is placed in communication with the product inside the receptacle by causing a part carrying the applicator to move along the longitudinal axis of the receptacle. When the product is applied to an area to be treated, pressure is exerted on the applicator along this axis, so that the passage aperture tends to reclose as the product is being applied, which is not at all desirable.
Patent application Ser. No. FR 2 681 050 describes a device with a dispensing tip fitted with a cap in which screwing/unscrewing causes the tip to open/close. However, a dispensing tip of this kind is not suitable for the application of a nail varnish remover to remove the varnish from one nail. Moreover, unscrewing the cap causes the cap to move to the open position due to the presence of bosses on the sidewall of the tip. A configuration of this kind does not allow the cap to be screwed back onto the bottle, when the tip is closed by hand, owing to the presence of the bosses.
One of the objects of the invention is to provide a packaging and application device for a fluid product which does not have the drawbacks of the prior art.
A particular object of the invention is to provide a device which enables nail varnish to be removed from one nail without damaging the varnish on the other nails.
A further object of the invention is to provide a device which is easy to use and which enables the product and applicator to be packaged in a manner such that the applicator is isolated from the product contained in the receptacle.
A further object of the invention is also to provide a device of this kind wherein the applicator element can be changed without risk of leakage of the product from the receptacle.

Packaging and application device for a product, notably a nail varnish remover


The present invention provides a method and apparatus for increasing the performance of world-wide-web traffic over the Internet. A distributed network of specialized nodes of two types is dispersed around the Internet. A web client's requests are directed to a node of the first type chosen to be close to the client, and the client communicates with this node using a standard protocol such as HTTP. This first node receives the request, and communicates the request to a node of the second type chosen to be close to the request's ultimate destination (e.g., a web server capable of generating a response to the request.) The first node communicates the request to the second node using a different, specialized, protocol that has been designed for improved performance and specifically to reduce traffic volume and to reduce latency. The second node receives communication from the first node using this specialized protocol, converts it back to a standard protocol such as HTTP, and forwards the request to the destination computer or server. Responses from the destination to the client take the corresponding reverse route, and also are carried over a specialized protocol between the two nodes. In addition, these nodes can employ other techniques such as web caches that avoid or improve some communication steps. Thus, specialized, proprietary, or complex protocols and techniques can be quickly deployed to enhance web performance without requiring significant changes to the clients or servers.

Today, improving the performance of Internet communication is a major technological and commercial concern. Investment in improving the Internet network infrastructure is estimated to become a $1.3 trillion dollar industry by 2003 (Source: Nortel Industries press release Jan. 31, 2000). It has been estimated that many web users will not tolerate a delay in downloading a web page of more than about 8 seconds, and that the current value of e-commerce sales at risk because of slow download speeds is $4.35 billion per year. (Source: Zona Research report "The need for speed", abstract available http://www.zonaresearch.com/info/press/99-jun30.htm). In this climate there is pressing demand for ways to improve web performance, and no simple or obvious techniques are overlooked.
Standard Internet Protocols: HTTP and TCP
Two standard protocols used on Internet links, HTTP and TCP, impose a significant limitation Internet communication speed. HyperText Transport Protocol (HTTP) is the application-level network protocol used when a client requests web content from a web server, and used by the web server when it responds to such requests. Modern network communication is layered, which means that higher-level protocols build on top of lower-level protocols (which in turn may build on other protocols). HTTP is a high-level protocol which includes commands to request content, respond with content, negotiate the form in which content is sent, and so forth. It is generally carried over the lower-level protocol Transmission Control Protocol (TCP). TCP enables reliable end-to-end connectivity between two locations in the Internet, but does not interpret the content sent between these two locations in any way: it just carries a stream of bytes. TCP in turn is generally carried over the Internet Protocol (IP), which is a packet-oriented protocol that does not guarantee reliable delivery.
It is well-known that HTTP and TCP are far from optimal protocols for Internet communication. TCP was developed and deployed well before HTTP was invented, and was designed for bulk bi-directional data transfer. HTTP is characterized by short request messages and moderate-length response transactions, and very bursty traffic. That TCP is not an optimal protocol for carrying HTTP is extensively documented. The paper "Modeling the Performance of HTTP Over Several Transport Protocols" in IEEE/ACM Transactions on Networking, vol. 5, number 5, October 1997, by Heidemann, Obraczka, and Touch, is representative of research addressing these issues. They claim, for instance:                These mismatches between the needs of HTTP and the services provided by TCP contribute to increased latency for most web users. Fundamentally, TCP is optimized for large-scale bulk data transport, while HTTP often needs a light-weight, request-response protocol.The mismatches referred to here relate to a number of technical features of TCP, including those known as "three way handshake", "slow-start congestion avoidance", and "TIME_WAIT tear-down delays". The Heidemann, Obraczka and Touch paper discusses several such improved protocols, such as Transaction TCP (T/TCP) and Asynchronous Reliable Delivery Protocol (ARDP). Other defects in TCP as it relates to HTTP include the flow-control algorithm being used, which can lead to unnecessary traffic and delays in the event of noise or error on the network.
HTTP itself is an evolving, improving protocol, but it has recognized performance deficiencies even aside from the interrelationship with TCP. The PhD Dissertation "Addressing the Challenges of Web Data Transport" by V. N. Padmanabhan (Computer Science Division, University of California at Berkeley, USA; Also published as Technical Report UCB/CSD-98-1016 September 1998) discusses some of these. As an example, it explains how HTTP Version 1.0 (still in wide use today) requires a client to send one request at a time over a given connection, waiting for the response to arrive completely before continuing, and at considerable performance cost (as the dissertation proves).
The problem is not that protocols for Internet communication that are better than HTTP and TCP do not exist or are not available. The problem is that HTTP and TCP are standards—widely accepted and widely deployed. Indeed, this is necessarily so, since communication over a shared network such as the Internet requires all users to use the same protocol. Thus, even when problems with existing protocols are noticed and improved protocols developed, it often takes a long time before such improvements become widely deployed. The delays are particularly long before improvements reach the public Internet infrastructure. In part, this delay is simply because costs are always large when significant software upgrades are needed. But in the case of protocol upgrades the costs and delays are even larger because no one can upgrade unilaterally: both ends of a network conversation must be using the same version of the same protocol. In the case of the Internet, some protocol changes require a community-wide coordinated update. For an example of such delays, consider that the problem with HTTP Version 1.0 cited above from Padmanabhan's dissertation was corrected in the next version of the protocol, where a feature known as "pipelining" was defined. Even several years after this improvement was first suggested there are very few web browsers that adopt it. Similarly, all proposals to replace TCP have languished, and today all major web browsers and web servers support HTTP over TCP only.
One prior art approach to improving Internet performance, without altering the standard protocols, is web caching. A similar approach is content distribution (CD). A Content Distribution (CD) network is a collection of specialized nodes or devices, placed in a larger network such the Internet at chosen locations such as in the offices of Internet Service Providers (ISPs). These nodes store certain web content on behalf of the content distributors' customers. Such stores are sometimes called caches, mirrors, or repeaters.
A Content Distribution service includes a redirection or interception service. When a web user (using a client such as a browser) requests content from a site, and the content is known or suspected to be cached at one or more CD nodes, the request is directed (or comes to be redirected) to some CD node that is "close" to the user. The notion of closeness is a measure of communications performance, and in particular can use such metrics as bandwidth capacity, bandwidth cost, latency, security, administrative boundaries, administrative convenience, and current congestion on various network paths. The technologies for choosing a close CD node and then directing requests to the chosen node are varied, but the field is still new and there is still considerable ongoing innovation.
An alternative to redirection of the type just discussed is interception, in which a node is placed in the network path from the client in such a way that it gets to see all web traffic from the client. A web proxy or other specialized device such as a router, for instance located at the client's ISP, can be used for this purpose. In this case, the node intercepts all traffic and if it sees a request for content it has cached (or can readily fetch from a nearby cache) then it can return the content immediately, but otherwise it relays the traffic to its destination unchanged. The use of a proxy may be under the client's control (e.g., if the client must be configured to use a proxy), or be "transparent" if the client needs no such configuration.
The advantages of Content Distribution are the possibility of serving traffic to the user from a close CD node, thus getting the response to him faster, cheaper, with less bandwidth, and perhaps more reliably. It is common to see reports of up to 10× improvement in the speed at which content is served to the end user.
The major disadvantage of Content Distribution is that not all content is effectively cacheable. It is particularly inappropriate for dynamically generated content, but also ineffective for rapidly changing content and some rarely accessed content. A CD node typically stores images, video files, sound files, static text pages, and other such content which does not change much from user to user. Such content is kept on the CD server in anticipation of requests for it (or perhaps, if there has already been one request, in anticipation of additional requests). However, much content on the Internet is generated on-the-fly in response to a customer's request; for instance, generated by a server program using the Common Gateway Interface (i.e. A "cgi-bin" program). Since the output of such a program may never be the same twice, or at least be likely to differ from person to person and from occasion to occasion, it is generally not feasible to have such content prepared in advance. There are, after all, hundreds of millions of web users; one could not generate and store this many customized pages in advance. As web content becomes more personal and more customized to each user, the importance of such pages will increase further. CD networks cannot anticipate such pages and so generally cannot improve the speed at which they are served.
A second disadvantage of Content Distribution arises because even so-called "static" content, such as images and fixed text, may be subject to occasional change. It is important to ensure that the caching node or Content Distribution node do not serve "stale" content, i.e. content that is no longer in agreement with the definitive copy on the origin server. A variety of schemes are used to ensure that content is fresh, or to lower the probability of delivering stale content. The mature field of caching technology addresses such issues. However, by the nature of the problem there is no perfect solution to this problem. To illustrate the issues, consider that the most recent version of the web protocol, HyperText Transfer Protocol Version 1.1, includes support for caching and Content Distribution that works as follows. A node with a cache can send a short message to the origin server asking, in effect, whether the copy of a web object held by the cache is still up to date. If so, a short acknowledgment is returned to the cache. If the cache node or CD node always makes such an inquiry before delivering content to a client then there is no chance of delivering stale content. But there is a delay, possibly large, as the message is sent to the origin server and the response is received. This scheme may reduce the volume of traffic sent over the network (bandwidth consumption) but does not necessarily reduce the delay before the content is seen by the client (latency). Such tradeoffs are inherent to any caching or Content Distribution technology.
A third disadvantage of caching and Content Distribution technology is that it requires significant computer resources, since a cache keeps copies of web content just in case a client will request them. A cache may keep many objects that are not, in fact, ever requested by a client before they become stale, and these consume expensive resources such as memory or disk space. The problem is made worse by the fact that a typical Content Distribution network has numerous caching nodes. There are many techniques that alleviate this problem somewhat, e.g. by using advanced algorithms to carefully distribute cached content across a network of multiple caching nodes. However, the high resource requirement is mostly inherent to the technology and can only be reduced, but not eliminated, by such techniques.
The term "caching" (or "proxy caching") is sometimes used to refer to a technique related to Content Distribution. There are only slight technical differences. "Caching" is more often heard when interception technology is used rather than redirection technology. Second, nodes are more likely to be called caches if they are operated on behalf of the clients rather than on behalf of the content originators. A related technology is server-side caching (also known as "reverse proxying") in which a cache node is located near the server rather than near clients. This technology sometimes delivers smaller performance gains than conventional caching or Content Distribution, but can often be deployed at reduced resource cost because only one such node is needed.
All forms of caching share the first two of the disadvantages of Content Distribution described above, the most critical being the inability to handle dynamically generated content.
There is therefore a need in the art for an approach to improving the performance of Internet communication, particularly communication between web clients and web servers, which does not require significant computer resources and which is compatible with existing standard protocols.

This patent is subject to a terminal disclaimer.

Method for high-performance delivery of web content


A small decoration capable of exhibiting play characteristics sufficient to provide a key ring with unexpectedness. The small decoration includes a container provided with a key ring mounting section. In the container is arranged a spring drive unit including a drive mechanism which includes a plurality of gears driven by a spiral spring. The drive mechanism also includes two gear shafts arranged so as to vertically project from the spring drive unit. One of the gear shafts is mounted with a decorative element such as a doll and the other gear shaft is mounted with a spring winding-up element. The spring winding-up element is adapted to wind up the spiral spring in association with closing of a lid member pivotally connected to the container and be returned to the original state while unwinding the spiral spring to actuate the decorative element when the lid member is open.

This invention relates to a small decoration adapted to be used while being mounted on personal belongings such as, for example, a key ring or the like, and more particularly to a small decoration equipped with a spring-operated movable decorative element constructed so as to exhibit play characteristics.
A key holder of one type which is one example of personal belongings generally includes a key ring threaded through a key to hold it thereon and a decorative element mounted on the key ring. The decorative element may include a plate made of a synthetic resin material, a doll, or the like. A key holder of another type which is conventionally known in the art is formed into a shape like a change purse and is provided with a key holding fitting adapted to hold a plurality of keys thereon.
Unfortunately, the conventional personal belongings including such a key ring as described above are not constructed so as to exhibit play characteristics, to thereby fail to give a user or viewer pleasure.

Small decoration equipped with spring-operated movable decorative element


The present invention discloses methods and compositions comprising a group of novel expression cassettes which provide significantly improved levels of accumulation of Coleopteran inhibitory Cry3B and Cry3B variant amino acid sequences when these are expressed in plants. The preferred embodiments of the invention provide at least up to ten fold higher levels of insect controlling protein relative to the highest levels obtained using prior compositions. In particular, transgenic maize expressing higher levels of a protein designed to exhibit increased toxicity toward Coleopteran pests deliver superior levels of insect protection and are less likely to sponsor development of populations of target insects that are resistant to the insecticidally active protein.

1. Field of the Invention
The present invention discloses transgenic plants expressing substantially higher levels of insect controlling Bacillus thuringiensis xcex4-endotoxin. Methods for obtaining such plants and compositions, and methods for using such plants and compositions are described. Also disclosed are improved polynucleotide cassettes containing preferred protein coding sequences which impart the substantially higher levels of insect controlling xcex4-endotoxins. The preferred embodiments of the invention surprisingly provide up to ten fold higher levels of insect controlling protein relative to the highest levels obtained using prior compositions. In particular, transgenic maize expressing higher levels of a protein designed to exhibit increased toxicity toward Coleopteran pests deliver superior levels of insect protection and are less likely to sponsor development of populations of target insects that are resistant to the insecticidally active protein.
2. Description of the Related Art
Almost all field crops, plants, and commercial farming areas are susceptible to attack by one or more insect pests. Particularly problematic are Coleopteran and Lepidopteran pests. Because crops of commercial interest are often the target of insect attack, environmentally-sensitive methods for controlling or eradicating insect infestation are desirable. This is particularly true for farmers, nurserymen, growers, and commercial and residential areas which seek to control insect populations using ecologically friendly compositions.
The most widely used environmentally-sensitive insecticidal formulations developed in recent years have been composed of microbial protein pesticides derived from the bacterium Bacillus thuringiensis, a Gram-positive bacterium that produces crystal proteins or inclusion bodies which are specifically toxic to certain orders and species of insects. Many different strains of B. thuringiensis have been identified which produce one or more insecticidal crystal proteins as well as other insecticidal non-crystal forming proteins. Compositions including B. thuringiensis strains which produce insecticidal proteins have been commercially available and used as environmentally acceptable insecticides because they are quite toxic to specific target insect pests, but are harmless to plants and to vertebrate and invertebrate animals. More importantly, because these insect controlling proteins have to be ingested by susceptible target insect pests in order to exert their insecticidal or toxic effects, judicious application of such protein compositions limits or prevents non-target insect members of the susceptible order which may also be susceptible to the composition from significant exposure to the proteins (for example, non-target Lepidopteran species where Lepidopteran specific B.t. crystal protein is used in an insecticidal formulation). Additionally, insects of various orders have been shown to totally lack susceptibility to specifically targeted insecticidal proteins even when ingested in large amounts.
xcex4-ENDOTOXINS
xcex4-endotoxins are used to control a wide range of plant-eating caterpillars and beetles, as well as mosquitoes. These proteins, also referred to as insecticidal crystal proteins, crystal proteins, and Bt toxins, represent a large collection of insecticidal proteins produced by B. thuringiensis that are toxic upon ingestion by a susceptible insect host. Over the past decade research on the structure and function of B. thuringiensis toxins has covered all of the major toxin categories, and while these toxins differ in specific structure and function, general similarities in the structure and function are assumed. A recent review describes the genetics, biochemistry, and molecular biology of Bt toxins (Schnepf et al., Bacillus thuringiensis and its Pesticidal Crystal Proteins, Microbiol. Mol. Biol. Rev. 62:775-806, 1998). Based on the accumulated knowledge of B. thuringiensis toxins, a generalized mode of action for B. thuringiensis toxins has been created and includes: ingestion by the insect, solubilization in the insect midgut (a combination stomach and small intestine), resistance to digestive enzymes sometimes with partial digestion by gut specific proteases catalyzing specifically a cleavage at a peptide site within a protoxin structure which xe2x80x9cactivatesxe2x80x9d the toxin, binding of the toxin to the midgut cells"" brush border, formation of a pore in the insect midgut cell, and the disruption of cellular homeostasis (English and Slatin, 1992).
GENES ENCODING CRYSTAL PROTEINS
Many of the xcex4-endotoxins are related to various degrees by similarities in their amino acid sequences. Historically, the proteins and the genes which encode them were classified based largely upon their spectrum of insecticidal activity. A review by Hxc3x6fte and Whiteley (1989) discusses the genes and proteins that were identified in B. thuringiensis prior to 1990, and sets forth the nomenclature and classification scheme which has traditionally been applied to B. thuringiensis genes and proteins. The original nomenclature took advantage of the discovery that the few Bt Cry proteins known at the time generally fell into a limited number of classes, wherein each class represented proteins having specificity for specific orders of insects. For example, cry1 genes encoded Lepidopteran-toxic Cry1 proteins. cry2 genes encoded Cry2 proteins that were generally toxic to both Lepidopterans as well as to Dipterans. cry3 genes encoded Coleopteran-toxic Cry3 proteins, while cry4 genes encoded Dipteran-specific toxic Cry4 proteins. The nomenclature has, for the past decade or more become rather confusing with the discovery of more distantly related classes of insecticidal Bt proteins. More recently, a simplified homogeneous nomenclature and basis for classifications of Bt proteins has been adopted and has been reviewed by Schnepf et al. (1998). Schnepf et al. (1998) also provides a structural solution for a Cry1 crystal. This simplified nomenclature will be adopted herein. The convention of identifying Bt genes with lower case, italicized letters (eg. cry1Ab1) and identifying Bt proteins with uppercase first character (eg. Cry1Ab1) will also be observed herein.
Based on the degree of sequence similarity, the proteins have been further classified into subfamilies. Proteins which appeared to be more closely related within each family were assigned divisional letters such as Cry1A, Cry1B, Cry1C, etc. Even more closely related proteins within each division were given names such as Cry1Ca, Cry1Cb, etc. and still even more closely related proteins within each division were designated with names such as Cry1Bb1, Cry1Bb2, etc.
The modern nomenclature systematically classifies the Cry proteins based upon amino acid sequence homology rather than upon insect target specificities. The classification scheme for many known toxins, not including allelic variations in individual proteins, is summarized in regularly updated tables which can be obtained from Dr. Neil Crickmore at at the biology department of Sussex University in Great Britain.
BIO-INSECTICIDE POLYPEPTIDE COMPOSITIONS
The utility of bacterial crystal proteins as insecticides was extended beyond Lepidopterans and Dipteran larvae when the first isolation of a Coleopteran-toxic B. thuringiensis strain was reported (Krieg et al., 1983; 1984). This strain (described in U.S. Pat. No. 4,766,203, specifically incorporated herein by reference), designated B. thuringiensis var. tenebrionis, was reported to be toxic to larvae of the Coleopteran insects Agelastica alni (blue alder leaf beetle) and Leptinotarsa decemlineata (Colorado potato beetle).
U.S. Pat. No. 5,024, 837 also describes hybrid B. thuringiensis var. kurstaki strains which showed activity against Lepidopteran insects. U.S. Pat. No. 4,797,279 (corresponding to EP 0221024) discloses a hybrid B. thuringiensis containing a plasmid from B. thuringiensis var. kurstaki encoding a Lepidopteran-toxic crystal protein-encoding gene and a plasmid from B. thuringiensis tenebrionis encoding a Coleopteran-toxic crystal protein-encoding gene. The hybrid B. thuringiensis strain produces crystal proteins characteristic of those made by both B. thuringiensis kurstaki and B. thuringiensis tenebrionis. U.S. Pat. No. 4,910,016 (corresponding to EP 0303379) discloses a B. thuringiensis isolate identified as B. thuringiensis MT 104 which has insecticidal activity against Coleopterans and Lepidopterans. More recently, Osman et al. disclosed a natural Bacillus thuringiensis isolate which displayed activity against at least two orders of insects and against nematodes (WO 98/30700).
It has been known for more than two decades that compositions comprising Bt insecticidal proteins are effective in providing protection from insect infestation to plants treated with such compositions. More recently, molecular genetic techniques have enabled the expression of Bt insecticidal proteins from nucleotide sequences stably inserted into plant genomes (Perlak et al., Brown and Santino, etc.). However, expression of transgenes in plants has provided an avenue for increased insect resistance to Bt""s produced in plants because plants have not been shown to produce high levels of insecticidal proteins. It was initially believed that gross morphological or topological differences in gene structure and architecture between plant and bacterial systems was the limitation which prevented over-expression of Bt transgenes in plants. These differences were seemingly overcome as disclosed by Perlak et al. (U.S. Pat. No. 5,500,365) and by Brown et al. (U.S. Pat. Nos. 5,424,412 and 5,689,052) wherein transgenes encoding Bt insecticidal protein which contained plant preferred codons were shown to improve the levels of expression. Alternatively, truncating the protoxin coding domain to the shortest peptide coding domain which still encoded an insecticidal protein was also deemed sufficient to overcome the limitation of vanishingly low expression levels of the Bt encoding transgene in planta. Expression levels of Bt proteins in planta from transgenes has varied widely independent of the means used for expression, and accumulated protein levels have ranged from virtually undetectable to 2 parts per million to around 20 to 30 parts per million. However, even though all of these approaches provided improved levels of Bt protein accumulation in plants, none provided levels of expression which could ensure that insect resistance would not become a problem without the necessity of coordinate expression of one or more additional insecticidal toxins by the transgenic plant, or alternatively without the coordinate topical application of additional supplemental Bt or insecticidal chemical compositions.
The importance of accumulation of higher levels of Bt toxin for preventing insect resistance to individual Bt toxins has been understood for some time. Various laboratory studies in which selection against Bt was applied over several generations of insects have confirmed that resistance against Bt insecticidal proteins is seldom obtained. It should be emphasized that laboratory conditions represent rather low but constant selection pressure conditions, allowing for the survival of a sub-population of insects which have been subjected to insecticidal pressure and which produce the subsequent generations of insects. Succeeding generations are also maintained on media containing low but constant concentrations of insecticidal protein. Generally, concentrations used for selection pressures range from LC40 to around LC60 or so, however, LC95 concentrations have also tested for the development of resistance. In most cases, resistance is acquired slowly, generally developing within a reasonably few generations, for example 10-50 generations. However, such resistance is not observed where substantially higher levels of toxin are used, or in situations in which multiple toxins are provided.
At present, recombinant plants expressing commercially useful levels of Bt insecticidal protein generally contain only one gene encoding a single class of Bt. Such plants are anticipated to have a very limited duration of use for two reasons. First, these plants are expressing insufficient levels of the insecticidal protein to ensure that all target insects exposed to and feeding from the plant tissues will succumb due to the dose of toxin ingested. Second, because of the insufficient insecticidal protein levels, the potential for development of resistance is unreasonably increased. This is not to say that the level of toxin produced by such transgenic plants is insufficient to be effective. This merely represents the limitations of expression of xcex4-endotoxins in planta even when using sequences encoding Bt xcex4-endotoxin which have been modified to conform to plant preferred sequences. One limitation which has been observed for many Bt xcex4-endotoxin encoding sequences modified for expression in plants is that is has been impossible to predict which Bt xcex4-endotoxin would be effective for expression in plants. (For example, expression of Cry2Aa in cotton plants results in phytotoxicity when targeted to the chloroplast, however expression of a closely related cry2Ab sequence is not phytotoxic when targeted to the chloroplast. (Corbin et al., U.S. patent application Ser. No. 09/186,002 ). Even so, levels of xcex4-endotoxin protein produced in plants is not sufficient to be effective against all desired target insect species known to be susceptible to a given type and class of xcex4-endotoxin.
As indicated above, alternative approaches to development of resistance to insecticidal proteins has included ineffective attempts to increase the expression levels of transgenes in plants. Alternatively, additional insecticidal genes could be engineered into plants so that multiple toxins are coordinately expressed. This would provide a more effective means for delaying the onset of resistance to any one combination of Bt""s, however, this still does not overcome the limitation of insufficient levels of insecticidal protein accumulating in the recombinant plant(s). An additional alternative to insufficient levels of expression has been to engineer genes encoding Bt insecticidal crystal proteins which demonstrate improved insecticidal properties, having either a broader host range or an increased biological activity, which could conceivably result in requiring less of the recombinant protein to control a target insect species than was required of the native form of the protein.
The combination of structural analyses of B. thuringiensis toxins followed by an investigation of the function of such structures, motifs, and the like has taught that specific regions of crystal protein endotoxins are, in a general way, responsible for particular functions.
Domain 1, for example, from Cry3Bb and Cry1Ac has been found to be responsible for ion channel activity, the initial step in formation of a pore (Walters et al., 1993; Von Tersch et al., 1994). Domains 2 and 3 have been found to be responsible for receptor binding and insecticidal specificity (Aronson et al., 1995; Caramori et al., 1991; Chen et al. 1993; de Maagd et al., 1996; Ge et al., 1991; Lee et al., 1992; Lee et al., 1995; Lu et al., 1994; Smedley and Ellar, 1996; Smith and Ellar, 1994; Rajamohan et al., 1995; Rajamohan et al., 1996; Wu and Dean, 1996). Regions in domain 2 and 3 can also impact the ion channel activity of some toxins (Chen et al., 1993, Wolfersberger et al., 1996; Von Tersch et al., 1994).
Unfortunately, while many investigators have attempted, few have succeeded in making mutated crystal proteins with improved insecticidal toxicity. In almost all of the examples of genetically-engineered B. thuringiensis toxins in the literature, the biological activity of the mutated crystal protein is no better than that of the wild-type protein, and in many cases, the activity is decreased or destroyed altogether (Almond and Dean, 1993; Aronson et al., 1995; Chen et al., 1993, Chen et al., 1995; Ge et al., 1991; Kwak et al., 1995; Lu et al., 1994; Rajamohan et al., 1995; Rajamohan et al., 1996; Smedley and Ellar, 1996; Smith and Ellar, 1994; Wolfersberger et al., 1996; Wu and Aronson, 1992). However, Van Rie et al. have recently accomplished the improvement of a Cry3A xcex4-endotoxin having increased Coleopteran insecticidal activity by identifying a single mutant having increased insecticidal activity. Van Rie et al. propose a method for identifying mutants having increased insecticidal activity in which the method consists of identifying amino acid mutations which decrease the insecticidal activity, and selectively altering those residues by site directed mutagenesis to incorporate one or more of the naturally occurring 20 amino acids at those positions, and feeding the various forms of the resulting altered protein to western or northern corn rootworms to identify those having improved activity (U.S. Pat. No. 5,659,123). While no sequences were enabled using the method, as mentioned above, Van Rie et al. succeeded in identifying only one sequence having increased activity and did not demonstrate an increase in expression of the mutant form as compared to the native sequence.
For a crystal protein having approximately 650 amino acids in the sequence of its active toxin, and the possibility of 20 different amino acids at each position in this sequence, the likelihood of arbitrarily creating a successful new structure is remote, even if a general function to a stretch of 250-300 amino acids can be assigned. Indeed, the above prior art with respect to crystal protein gene mutagenesis has been concerned primarily with studying the structure and function of the crystal proteins, using mutagenesis to perturb some step in the mode of action, rather than with engineering improved toxins.
Collectively, the limited successes in the art to develop non-naturally occurring toxins with improved insecticidal activity have stifled progress in this area and confounded the search for improved endotoxins or crystal proteins. Rather than following simple and predictable rules, the successful engineering of an improved crystal protein may involve different strategies, depending on the crystal protein being improved and the insect pests being targeted. Thus, the process is highly empirical.
Accordingly, traditional recombinant DNA technology is clearly not routine experimentation for providing improved insecticidal crystal proteins. What has been lacking in the prior art are rational methods for producing genetically-engineered B. thuringiensis crystal proteins that have improved insecticidal activity and, in particular, improved toxicity towards a wide range of Lepidopteran, Coleopteran, or Dipteran insect pests. Methods and compositions which address these concerns were disclosed in U.S. Pat. No. 6,063,597 (filed Dec. 18, 1997; English et al.) and other related U.S. Pat. No. 6,060,594, filed Dec. 18, 1997, English et al.; U.S. Pat. No. 6,077,824, filed Dec. 18, 1997, English et al.; and U.S. Pat. No. 6,023,013, filed Dec. 18, 1997, English et al.) and in Van Rie et al. (U.S. Pat. No. 5,659,123, Jun. 1, 1999). In addition, recombinantly improved xcex4-endotoxins have continued to be expressed poorly and/or cause phytoxic effects when expressed in plants, thus leading to the recovery of fewer commercially useful transgenic events.
Described herein are novel compositions and methods for expressing in transformed plants variant Cry3 B. thuringiensis xcex4-endotoxins having significant Coleopteran inhibitory activity. These compositions and methods advantageously result in plants expressing B. thuringiensis Cry3 67-endotoxins at increased levels not previously observed for Cry xcex4-endotoxins. Increased levels of Cry3 xcex4-endotoxin expression are reflected in the attainment of higher maximal expression levels in individual transgenic insertion events. Unexpectedly, the particular compositions disclosed herein result in the recovery of an increased percentage of transgenic events which manifest expression levels that far exceed threshold levels of expression necessary for Coleopteran insect control and which provide sufficient toxin levels capable of supporting a resistance management strategy. Since Cry3 xcex4-endotoxins are typically less potent than other xcex4-endotoxins commonly used to control Lepidopteran or Dipteran target pests when expressed in transgenic plants, attainment of higher maximal levels of Cry3 xcex4-endotoxin expression and recovery of more transgenic events with effective expression levels are both critical in isolating transgenic events expressing Cry3 xcex4-endotoxin which exhibit commercially useful levels of target insect control.
Another limitation of the prior art addressed by the present invention is the development of insect resistance to xcex4-endotoxins provided by plant expression. Specifically, the instant invention provides a superior strategy for the delay or elimination of the development of resistance to Cry3 xcex4-endotoxins through improved accumulation of xcex4-endotoxin within plant cells so that levels of the xcex4-endotoxin are maintained in-planta above a threshold level of protein, typically measured in parts per million (ppm). Improved expression of xcex4-endotoxins, which also should be taken to mean increased expression in view of what has been previously observed in the art, is believed to result in delayed onset of insect resistance and thus extends the utility of plant expressed xcex4-endotoxins as insect control agents.
In preferred embodiments, the present invention provides isolated and purified novel Cry3B xcex4-endotoxin proteins exhibiting particularly effective insecticidal activity directed toward controlling Coleopteran pest insect species. Such xcex4-endotoxin proteins of the present invention are provided by expression from isolated, purified and improved or enhanced DNA or polynucleotide sequences each comprising a Cry3 xcex4-endotoxin coding sequence placed under the control of preferred plant functional gene expression elements such as a promoter, an untranslated leader sequence, an intron and a transcription termination and polyadenylation sequence. Some preferred DNA or polynucleotide sequences may also provide for plastid or chloroplast targeting protein sequences. Preferred DNA constructs of the present invention include those constructs which encode Cry3 xcex4-endotoxins exhibiting Coleopteran-inhibitory or Coleopteran-controlling activity. In an illustrative embodiment, polynucleotide sequences are assembled into an expression cassette for introduction into plant genomic DNA, wherein the expression cassette comprises a Cry3Bb xcex4-endotoxin variant coding sequence operably linked to a sequence comprising a promoter, an untranslated leader sequence, an intron and a transcription termination and polyadenylation sequence. In particular, a transgene localized within a plant operable polynucleotide expression cassette or polynucleotide sequence comprising an expression cassette which is comprised of genetic elements which function in plant cells to express a desired protein from a nucleic acid coding sequence (the transgene) which is operably localized within said expression cassette. The coding sequence is linked upstream to at least a promoter sequence, an untranslated leader sequence (UTL), an intron sequence, and in-frame in certain indicated embodiments to a sequence encoding a plastid or chloroplast targeting peptide. The coding sequence is also linked downstream to at least a plant functional transcription termination and polyadenylation sequence. Polynucleotide sequences comprising such an expression cassette are shown herein to improve expression of the desired protein encoded from within the cassette, improve the number of events obtained from the use of the polynucleotide sequence in plant transformation, wherein said improved number of events contain the desired transgene localized within the expression cassette and exhibit improved levels of expression of one or more desired proteins. The improved number of events are also surprisingly observed to express the desired protein at levels above 2 to 5 parts per million but in general below 200 to 500 parts per million of total cell protein. Even more surprising were some events in particular which expressed the desired protein at levels well above 500 ppm. Indicated embodiments disclose a sequence encoding a variant Cry3Bb xcex4-endotoxin comprising the isolated and purified SEQ ID NO:9, from NcoI to EcoRI as set forth in FIG. 1 illustrating plasmid pMON25096. Yet other embodiments disclose a variant Cry3Bb xcex4-endotoxin coding sequence comprising an isolated and purified SEQ ID NO:11, from NcoI to EcoRI as set forth in FIG. 2 illustrating plasmid pMON33741. It is contemplated, however, that any Cry3 xcex4-endotoxin exhibiting substantial Coleopteran-inhibitory or Coleopteran-controlling activity greater than or equal to that disclosed in the present invention could be utilized according to the embodiments of the present invention, with those Cry3 proteins bearing substantial homologies to Cry3Bb being particularly preferred.
In a preferred embodiment, the invention provides for transgenic plants which have been transformed with a DNA construct or expression cassette of the present invention that is expressed and translated at unexpectedly high levels by the plant which results in surprisingly high levels of xcex4-endotoxin accumulation. Monocotyledenous plants may be transformed according to the methods and with the DNA constructs disclosed herein. However, it is also anticipated that dicotyledenous plants could also be transformed with DNA sequences disclosed herein by one skilled in the art in order to obtain transgenic plants providing unexpectedly useful levels of insect resistance without the risk of development of insect resistance to the xcex4-endotoxin. The plant transformed by the instant invention may be prepared, in a further preferred embodiment, by a process including obtainment of the isolated and purified DNA construct contained within the expression cassette, and then transforming the plant with the construct so that the plant expresses the protein for which the construct encodes. Alternatively, the plant transformed by the instant invention may be prepared, in a further preferred embodiment, by a process including introduction of the isolated and purified DNA construct into a transformation competent Agrobacterium strain, and then transforming the plant with the Agrobacterium strain containing the construct so that the plant expresses the proteins for which the construct encodes. It has been observed herein that transformation of plants by the disclosed compositions and methods results surprisingly in increased frequencies of transformants exhibiting transgene expression as well as in the recovery of individual transgenic events exhibiting unexpectedly higher absolute levels of transgene expression.
It is contemplated that the increased expression levels observed in the disclosed invention will allow for reduced development of insect resistance to Bt xcex4-endotoxins presented to target insect pests. This may be achieved by transforming a plant with the preferred DNA construct to achieve high rates of Cry3 expression alone, or by simultaneously exposing target insects to the disclosed Cry3 xcex4-endotoxins along with other compositions effective in controlling Coleopteran species such as variants of Cry3B (English et al., WO 99/31248), variant Cry3A or variant Cry3D (U.S. Pat. No. 5,659,123), CryET33 and CryET34 (Donovan et al., WO 97/17600), CryET70 (U.S. application Ser. No. 09/184,748; Mettus et al., Nov. 2, 1998), Cry6A, Cry6B, Cry8B (U.S. Pat. No. 5,277,905), CryET29 (Rupar et al., WO 97/21587), insecticidal acyl lipid hydrolases, combinations of amino acid oxidases and tedanalactam synthases (Romano et al., U.S. application Ser. No. 09/063,733, filed Apr. 21, 1998), or insecticidal proteins such as VIP1 (Gay, WO 97/26339; Gourlet et al., WO 98/02453) and VIP3 (Estruch et al., U.S. Pat. No. 5,877,012; 1999) among others. Susceptible target insects include Diabroticus spp. Wire Worm in Zea mays and Leptinotarsa decemlineata (Say) in Solanum tuberosum, and Boll Weevil in Gossypium species (cotton).
It is therefore contemplated that the compositions and methods disclosed by the present invention will provide many advantages over the prior art including those specifically outlined above. Other advantages include improved control of susceptible target insect pests and achieving season long protection from insect pathogens. An additional advantage of the present invention provides for reducing the number of transgenic events that have to be screened in order to identify one which contains beneficial levels of one or more insect controlling compositions. The present invention also encompasses cells transformed with the DNA constructs disclosed herein. Also, transformation vectors such as plasmids, bacmids, artificial chromosomes, viral vectors and such are contemplated as elements for use in delivering the nucleotide compositions of the present invention into contemplated cells in order to obtain transformed host cells, both prokaryotic and eukaryotic, which express the xcex4-endotoxin proteins encoded by the novel DNA construct disclosed herein. It is further contemplated that in some instances the genome of a transgenic plant of the present invention will have been augmented through the stable integration of an expression cassette encoding a Coleopteran inhibitory or controlling B. thuringiensis xcex4-endotoxin or variants thereof as described herein. Furthermore, more than one transgene encoding an insecticidal composition will be incorporated into the nuclear genome, or alternatively, into the chloroplast or plastid genome of the transformed host plant cell. It is envisioned that more than one polynucleotide encoding an insecticidal crystal protein will be incorporated into the genome of a plant cell and it may be desirable to have two or even more sequences encoding insecticidal or other plant beneficial proteins within the nucleotide sequences contained within the cell. Such recombinantly derived proteins may exist as precursors, pro-toxins, or as fusions of beneficial proteins linked by flexible amino acid linker sequences or by protease specific cleavage sequences well known in the art. Chimeras comprising fusions of insecticidal proteins are also envisioned. The offspring of transgenic plant host cells can be manipulated artificially to produce whole recombinant plants exhibiting improved insecticidal properties, and the recombinant nucleotide sequences are shown herein to be heritable. The heritability of the elements is a preferred aspect of this invention, so that the expression elements are able to be delivered to lineal descendants of the original transformed host plant cell, giving rise first to a stably transformed plant whose constituent cells express the desired transgene, albeit tissue specific expression can be selectively manipulated generally through the choice of plant operable promoter selected for use in a given expression cassette, as described above. Transformed plants give rise to seeds containing the heritable expression cassette, and the seeds thus give rise to plants in lineal fashion which contain the expression cassette, generally in Mendelian fashion, particularly when selfed according to well known methods in the art.

Expression of Cry3B insecticidal protein in plants


Ablation instruments and techniques are disclosed for accessing portions of the heart wall via the coronary sinus. While shielding the major portion of the sinus and/or the circumflex coronary artery from ablative energy, the present invention provides access to the heart wall to treat atrial fibrillation. In particular, the present invention provides instruments and methods for forming transmural lesions in the left atrium, such as the so-called “left isthmus block,” from within the coronary sinus.

Atrial fibrillation (AF) is a form of cardiac arrhythmia characterized by rapid randomized contractions of atrial myocardium, causing an irregular, often rapid ventricular rate. The regular pumping function of the atria is replaced by a disorganized, ineffective quivering. Atrial fibrillation is caused, in large part, by chaotic conduction of electrical signals through the upper chambers of the heart. Atrial fibrillation is often associated with other forms of cardiovascular disease, including congestive heart failure, rheumatic heart disease, coronary artery disease, left ventricular hypertrophy, cardiomyopathy or hypertension.
Various techniques have been proposed for the treatment of atrial fibrillation by blocking errant electrical signal conduction in the heart. For example, the "Maze procedure" was developed in the early 1990s to treat atrial fibrillation by forming a series of physical incisions (in the form of a maze) to create scar tissue in the heart that would stop or redirect the electrical impulses. Although these procedures were originally performed with a scalpel, various other techniques have also been developed to form lesions.
Researchers have come to realize that the origins of AF often lie in the left atrium in the vicinity of the pulmonary veins. A lesion encircling the pulmonary veins is currently considered to be one of the most important areas to isolate. Additionally, a second lesion extending from the pulmonary veins to the mitral valve annulus is also considered advantageous. This mitral valve annulus lesion is sometimes referred to as the "left isthmus block."
Typically, such lesions are formed from within the heart (by applying a scalpel or other ablation instrument to the endocardial tissue). In fact, it is becoming increasingly commonplace for patients undergoing open heart cardiac surgery (e.g., for mitral valve replacement) to also have an ablative procedure if they have a history of AF.
These procedures are very invasive (requiring a sternotomy, cardiopulmonary bypass, cardioplegic arrest, and cardiac incisions) to enter the left atrium of the heart and create the lesions. Despite various advances in rapid lesion-forming devices (such as ultrasound, radio-frequency (RF), laser and cryogenic instruments), open heart surgery has a significant morbidity and remains a procedure of last resort for treating AF except when other conditions mandate such surgery.
Considerable effort has been made to find ways to treat atrial fibrillation without cardiac arrest and open-heart surgery. For example, light based systems for creating lesions on the outer surface of the heart have been disclosed in U.S. Pat. No. 6,558,375 entitled "Surgical Ablation Instruments for Forming an Encircling Lesion" herein incorporated by reference in its entirety. In such systems, radiant energy is applied to the epicardium to induce a transmural lesion. These systems permit the surgeon to encircle the pulmonary veins with an instrument and create a lesion by photoablation. Others have proposed similar epicardial devices based on ultrasound and the like. Such systems avoid the need to stop the heart (cardioplegic arrest).
Although epicardial ablation devices have been proposed to create lesions around the pulmonary veins, access to the regions of the atrium extending from the pulmonary veins to the mitral valve annulus remains a problem, primarily because other cardiac structures can preclude epicardial access. In particular, the coronary sinus and circumflex artery lie directly above the target region for creation of the left isthmus block or lesion. If ablative energy is applied to the circumflex coronary artery in an effort to form an atrial lesion, stenosis of this blood vessel can occur and potentially result in a myocardial infarction or even death.
Thus, there exists a need for better ablation instruments and techniques. In particular, instruments capable of forming lesions extending from the pulmonary veins to the mitral valve annulus would satisfy an important need in the field.

Ablative treatment of atrial fibrillation via the coronary sinus


A two stage process useful for cetane upgrading of diesel fuels. More particularly, the invention relates to a process for selective naphthenic ring-opening utilizing an extremely low acidic distillate selective catalyst having highly dispersed Pt. The process is a two stage process wherein the first stage is a hydrotreating stage for removing sulfur from the feed and the second stage is the selective ring-opening stage.

1. Field of the Invention
The present invention relates to a two stage process useful for cetane upgrading of diesel fuels. More particularly, the invention relates to a process for selective naphthenic ring-opening utilizing an extremely low acidic distillate selective catalyst having highly dispersed Pt. The process is a two stage process wherein the first stage is a hydrotreating stage for removing sulfur from the feed and the second stage is the selective ring-opening stage.
2. Description of Prior Art
Under present conditions, petroleum refineries are finding it increasingly necessary to seek the most cost-effective means of improving the quality of diesel fuel products. Cetane number is a measure of ignition quality of diesel fuels. Cetane number is highly dependent on the paraffinicity of molecular structures whether they be straight chain or alkyl attachments to rings. Distillate aromatic content is inversely proportional to cetane number while a high paraffinic content is directly proportional to a high cetane number.
Currently, diesel fuels have a minimum cetane number of 45. But the European Union (EU) just passed an amendment requiring that the cetane number of European diesel fuels reach 51 by the year 2000, even higher cetane numbers of at least 58 are being proposed for the year 2005 and beyond.
Aromatic compounds are a high source of octane, but they are poor for high cetane numbers. Aromatic saturation, which can be described as the hydrogenation of aromatic compounds to naphthene rings, has been commonly used to upgrade the cetane level of diesel fuels. However, aromatic saturation can only make low cetane naphthenic species, not high cetane components such as normal paraffins and iso paraffins. As a result, the use of a hydrocracking catalyst for the ring-opening of naphthenic species had been used to solve this problem.
Conventional hydrocracking catalysts that open naphthenic rings rely on high acidity to catalyze this reaction. Because hydrocracking with a highly acidic catalyst breaks both carbon-carbon and carbon-hydrogen bonds, the use of such a catalyst cannot be selective in just opening rings of naphthenic species without cracking desired paraffins for the diesel product.
Furthermore, commercial hydrocracking catalysts rely on acidity as the active ring-opening site, and this active site also catalyzes increased hydroisomerization of the resulting naphthenes and paraffins. It is typical for a cumulative loss of 18-20 cetane numbers for each methyl branching increase. The use of a low acidic catalyst would minimize diesel yield loss, the production of isoparaffins, and the production of gaseous by-products.
Hydroprocessing can be done in a co-current, counter-current or an ebullated bed configuration. In a conventional co-current catalytic hydroprocessing, a hydrocarbon feed is initially hydrotreated to help get rid of heteroatom-containing impurities. These heteroatoms, principally nitrogen and sulfur, are converted by hydrodenitrogenation and hydrodesulfurization reactions from organic compounds to their inorganic forms (H2S and NH3). These inorganic gases inhibit the activity and performance of hydroprocessing catalysts through competitive adsorption on the catalyst. Therefore, the catalyst containing portion of a conventional co-current reactor is often limited in reactivity because of low H2 pressure and the presence of high concentrations of heteroatom components.
Conventional counter-current configurations utilizes a device that creates a flow of hydrogen containing gas within a container in order to force the gaseous phase to flow counter to the liquid phase. U.S. Pat. No. 5,888,376 discloses a counter current process for converting light oil to jet fuel by first hydrotreating the light oil and then flowing the product stream counter-current to upflowing hydrogen containing gas in the presence of hydroisomerization catalysts. These hydroisomerizaton catalysts are highly acidic catalysts. U.S. Pat. No. 5,882,505 also discloses hydroisomerizing wax feedstocks to lubricants in a reaction zone containing an acidic hydroisomerization catalyst in the presence of a hydrogen-containing gas. U.S. Pat. No. 3,767,562 discloses making jet fuel by using a hydrogenation catalyst in a counter-current configuration. None of the counter-current methods in the prior art discloses the use of a catalyst that can selectively open naphthenic species without cracking desired paraffins.
In light of the disadvantages of the conventional processes for improving diesel fuel, there remains a need for a process of selective naphthenic ring-opening that produces an increased cetane number of diesel fuel without a corresponding diesel yield loss.
In accordance with the present invention, a process is provided for selective ring-opening of naphthenes catalyzed by a low acid catalyst in order to increase diesel fuel yield and cetane number.
In the process, a hydrocarbon feed is contacted with a hydrogen containing gas under superatmospheric conditions with a selective ring-opening (SRO) catalyst. Ideally, the process operates in a counter-current configuration in order to remove gaseous heteroatoms. In the counter-current configuration, the catalyst can operate at lower temperatures in order to minimize hydrocracking and hydroisomerization of paraffin, thereby increasing cetane number and diesel yield. The selective ring-opening catalyst preferably has a crystalline molecular sieve material component and a Group VIII noble metal component. The crystalline molecular sieve material component is a large pore faujasite structure having an alpha acidity of less than 1, preferably less than 0.3. Zeolite USY is the preferred crystalline molecular sieve material component.
The Group VIII noble metal component can be platinum, palladium, iridium, rhodium, or a combination thereof. Platinum is preferred. The content of Group VIII noble metal component can vary. The preferred range is between 0.1 and 5% by weight of the catalyst.
The Group VIII noble metal component is located within the dispersed clusters. In the preferred embodiment, the particle size of Group VIII metal on the catalyst is less than about 10 xc3x85. Dispersion of the metal can also be measured by hydrogen chemisorption techniques in terms of the H/metal ratio. In the preferred embodiment, when platinum is used as the noble metal component, the H/Pt ratio is between about 1.1 and 1.5.
The advantages of the present invention is that (1) it allows selective ring opening of naphthene rings by the use of a low acid catalyst in addition to hydrogenating aromatics and cracking heavy paraffins, and (2) it allows the low acid catalyst to operate at the lowest possible temperature by using a counter-current configuration in order to prevent undesired hydrocracking and hydroisomerization.
For a better understanding of the present invention, together with other and further advantages, reference is made to the following description, taken in conjunction with accompanying drawings, and its scope will be pointed out in the appended claims.

Selective ring opening process for producing diesel fuel with increased cetane number

ExxonMobil Research and Engineering Company


A method and system for aligning a sensor of a radar unit mounted on a vehicle, including calculating at least one alignment measurement of the vehicle from a plurality of optical sensors, positioning a target at a predetermined distance forward of the radar unit and determining a position of the radar unit based on the alignment measurement and the predetermined distance of the target. The method further includes transmitting a beam extending forward from the radar unit and positioning the target and the sensor in alignment with an axis of the beam.

Intelligent vehicle systems use sensor technology to detect objects surrounding a region of a host vehicle. For example, a collision avoidance system can include laser or radar sensors to detect possible collisions based on the movement of surrounding vehicles, objects and pedestrians. The collision avoidance system can control another vehicle system in response to the detection of an object. For example, a braking system or a steering system can be controlled to mitigate collision with the detected object. Proper performance of such intelligent vehicle systems requires precise alignment of the sensors to detect objects with a minimal error rate.

Vehicle radar alignment method and system


A method for producing carbon nanotubes, the method comprising: (a) providing a substrate with a top surface, (b) forming an island of catalyst material on the top surface using a tip having a patterning compound thereon, (c) heating the substrate and catalyst island, and (d) contacting the catalyst island with a carbon-containing gas for a period of time sufficient to form the nanotubes on the catalyst island.

New forms of carbon including carbon nanotubes have commercially attractive properties. Better, more commercially attractive methods are needed to produce such materials for nanotechnological advancement. In addition, better methods are needed to provide purer materials and to locate materials in existing structures. Location is vital, for example, to building nanoelectronic and nanooptical devices. Also, better methods are needed to produce single wall carbon nanotubes. Nanolithography provides a commercially attractive route to improve carbon nanotube technology and provide high resolution and high alignment capabilities.

Apparatus, materials, and methods for fabrication and catalysis


An apparatus and a method for controlling the shear rate at which an acoustic wave device measures viscosity, by utilizing an automatic level control or an automatic gain control circuit to control power input to the sensor as a function of the sensor's output power. Further improvement is provided by measuring the input power and combining the input power and output power measurements, preferably by averaging, to control the input power to the sensor. A method is also provided for characterizing the fluid under test by providing a set of viscosity measurements at various shear rates.

Viscosity, and more generally, viscoelasticity, are properties of liquids and solids that relate the shear forces generated by or applied to a material to the amount of shear deformation or flow. While the invention applies equally well to viscoelasticity, the present discussion will be limited to viscosity measurement for simplicity. Viscosity is of widespread interest in many manufacturing environments and is measured as a primary quality of some products and as a secondary quality (a means of monitoring process state) in other processes.
Viscosity describes the force required in order to make successive molecular layers of a liquid move past each other at a given rate of shear ("shear rate"). If one considers a liquid flowing past the walls of a container, the liquid will ideally have no motion relative to the wall at the interface and will have increasingly higher velocities as one observes points successively further from the wall. The shear rate is defined as the gradient of the velocity of the liquid parallel to the surface (meters per second) with increasing distance from the surface (meters). The units of shear rate are 1/seconds. The shear stress is the amount of force per unit area that must be applied in order to cause the motion. While the fluid may have a characteristic flow (and thus a characteristic shear rate) or may be stationary, all measurements of viscosity to date are based on the measurement of shear stress vs. shear rate under an imposed motion of the fluid. Throughout this disclosure, "shear rate at which the viscosity of a fluid" should be taken to mean the shear rate at which the viscosity of the fluid is measured, which may differ substantially from the characteristic shear rate of the fluid in its intended application or point of measurement.
In U.S. Pat. No. 7,007,546 the present inventor detailed a method for measuring the viscosity of a fluid at a selected shear rate, and methods for characterizing characteristics of a fluid by measuring the viscosity of the fluid at selected shear rates. This is achieved by utilizing an Acoustic Wave Device (AWD). The method for measuring viscosity at a desired shear rate utilizes an AWD sensor and an estimate of the shear rate at which the sensor operates, and then modifying the input power level to obtain the desired shear rate. It utilizes measurements of both the input and output power levels and the selection of an input power level by an algorithm. The method of characterizing the fluid characteristics include feeding different levels of power to the sensor and measuring fluid viscosities at the differing power levels. The present invention represents an improvement to the method described in the '546 patent.

Control of equivalent shear rate in acoustic wave sensors

Vectron International, a division of Dover Electronics, Inc.


Disclosed is a seat reclining mechanism, which is designed to be mounted to a seat cushion and a seat back so as to allow the seat back to be reclined. The seat reclining mechanism comprises a guide bracket and an internal gear member coupled to the guide bracket in a rotatable manner about a rotation axis thereof with respect to the guide bracket and formed with internal teeth along a circle having a center on the rotation axis. A plurality of lock gear members is disposed between the guide bracket and the internal gear member. A fitting portion is provided in either one of the guide bracket and the internal gear member. A rotatable cam is rotatably supported by the fitting portion.

1. Field of the Invention
The present invention relates to a seat reclining mechanism.
2. Description of the Related Art
Heretofore, there has been known a seat reclining mechanism designed to be mounted to a seat cushion and a seat back so as to allow the seat back to be reclined. JP 2001-017259A and JP 2004-154585A disclose a reclining mechanism which comprises a guide bracket adapted to be mounted to either one of a seat cushion and a seat back, an internal gear member (i.e., internally-toothed gear member) adapted to be mounted to a remaining one of the seat cushion and the seat back, a plurality of lock gear members disposed between the guide bracket and the internal gear member and adapted to be slidingly guided in a radial direction, a rotatable cam adapted to move the lock gear members in both a lock direction causing the lock gear members to be brought into engagement with the internal gear member and an unlock direction causing the lock gear members to be released from the engagement with the internal gear member, and a lock spring biasing the rotatable cam in a lock-causing rotation direction.
In this mechanism, when an operator operates an operation lever to rotate the rotatable cam in a direction opposite to the lock-causing rotation direction, the lock gear members are moved in the unlock direction through a circular lever (actuating plate), and released from the engagement with the internal gear member. Thus, the seat back can be forwardly or rearwardly inclined. Then, when the operator releases his/her hand from the operation lever, the rotatable cam is rotated in the lock-causing rotation direction by a biasing force of the lock spring, and the lock gear members are moved in the lock direction and brought into engagement with the internal gear member. Thus, the seat back is locked at an intended inclined position.
In the above mechanism, if a center of the rotatable cam is displaced from a normal position due to the biasing force of the lock spring, the lock gear members are likely to become unable to be evenly moved, which leads to instability in lock function of the lock gear members.
For this reason, the rotatable cam is integrated with an operation member, such as an operation rod connected to the operation lever, and the operation member supported by a bearing member, such as a knuckle bracket, provided outside the reclining mechanism, so as to perform centering of the rotatable cam.
However, in the structure designed to support the operation member by the bearing member, such as a knuckle bracket, the operation member and the bearing member have to be prepared with a high degree of dimensional accuracy to cause a problem about an increase in cost.