Abstract: Disclosed is a process of screening clones having DNA from an uncultivated microorganism for a specified protein, e.g. enzyme, activity by screening for a specified protein, e.g. enzyme, activity in a library of clones prepared by (i) recovering DNA from a DNA population derived from at least one uncultivated microorganism; and (ii) transforming a host with recovered DNA to produce a library of clones which is screened for the specified protein, e.g. enzyme, activity.

Abstract: Disclosed is a process for identifying compounds having a specified activity of interest, which process comprises (i) introducing interacting molecules into a host cell under conditions to generate or repress a detectable signal; and (ii) introducing a third compound or gene or genes encoding a third compound into the host cell from (i); and (iii) screening said host cell utilizing a method for detecting the inhibition or enhancement of interaction of proteins or other molecules in an in vivo or in vitro system. Another aspect of the present invention provides a process for identifying compounds of interest, which process comprises (i) generating one or more expression libraries derived from nucleic acid directly isolated from the environment; and (ii) screening said libraries utilizing a method for detecting the inhibition or enhancement of interaction of proteins or other molecules in an in vivo or in vitro system.

Abstract: Disclosed is a process for identifying clones having a specified activity of interest, which process comprises (i) generating one or more expression libraries derived from nucleic acid directly isolated from the environment; and (ii) screening said libraries utilizing a fluorescence activated cell sorter to identify said clones. More particularly, this is a process for identifying clones having a specified activity of interest by (i) generating one or more expression libraries derived from nucleic acid directly or indirectly isolated from the environment; (ii) exposing said libraries to a particular substrate or substrates of interest; and (iii) screening said exposed libraries utilizing a fluorescence activated cell sorter to identify clones which react with the substrate or substrates.

Abstract: A system and method for optically detecting samples held in a solution requires the use of a holding plate that has as many as one-thousand through-hole wells, or more. The solution is suspended in these through-hole wells under surface tension between opposed surfaces of the holding plate. A pneumatic pump is then engaged with the plate to establish a differential pressure (&Dgr;p) between the upper and lower surfaces of the solution that is equal to approximately two tenths of a pound per square inch (0.2 psi). The result is the formation of a convex meniscus on a surface of the solution that causes light passing into the solution to converge and concentrate. This concentration of light, in turn, facilitates optical detection of samples in the solution.

Abstract: Provided are methods of screening and identification of bio activities and bioactive molecules of interest using a capillary array system. More specifically, disclosed are methods of using optical detection and capillary array-based techniques for screening libraries and recovering bioactive molecules having a desired activity or a nucleic acid sequence encoding such bioactive molecules.

Abstract: Disclosed is a process for obtaining an enzyme having a specified enzyme activity derived from a heterogeneous DNA population by screening, for the specified enzyme activity, a library of clones containing DNA from the heterogeneous DNA population which have been exposed to directed mutagenesis towards production of the specified enzyme activity. Also disclosed is a process for obtaining an enzyme having a specified enzyme activity by screening, for the specified enzyme activity, a library of clones containing DNA from a pool of DNA populations which have been exposed to directed mutagenesis in an attempt to produce in the library of clones DNA encoding an enzyme having one or more desired characteristics which can be the same or different from the specified enzyme activity.

Abstract: This invention provides methods of obtaining novel polynucleotides and encoded polypeptides by the use of non-stochastic methods of directed evolution (DirectEvolution™). A particular advantage of end-selection-based methods is the ability to recover full-length polynucleotides from a library of progeny molecules generated by mutagenesis methods. These methods include non-stochastic polynucleotide site-saturation mutagenesis (Gene Site Saturation Mutagenesis™) and non-stochastic polynucleotide reassembly (GeneReassembly™). This invention provides methods of obtaining novel enzymes that have optimized physical &/or biological properties. Through use of the claimed methods, genetic vaccines, enzymes, small molecules, and other desirable molecules can be evolved towards desirable properties. For example, vaccine vectors can be obtained that exhibit increased efficacy for use as genetic vaccines.

Abstract: Disclosed is a rapid and facilitated method of producing from a parentlal template polynucleotide, a set of mutagenized progeny polynculeotides whereby at each original codon position there is produced at least one substitute codon encoding each of the 20 naturally encoded amino acids. Accordingly, there is also provided a method of producing from a parental template polypeptide, a set of mutagenized progeny polypeptides wherein each of the 20 naturally encoded amino acids is represented at each original amino acid position. The method provided is termed site-saturation mutagenesis, or simply saturation mutagenesis, and can be used in combination with other mutagenization processes, such as, for example, a process wherein two or more related polynucleotides are introduced into a suitable host cell such that a hybrid polynucleotide is generated by recombination and reductive reassortment.

Abstract: A holding plate for selectively heating and cooling samples in a solution has two opposing surfaces, and a plurality of cylindrically-shaped through-hole wells for holding the samples. Each well extends between the two surfaces of the holding plate, and has an aspect ratio of greater than 5:1, and a diameter less than approximately 500 microns. A metallic coating is applied by vapor deposition techniques on a surface of the holding plate. Importantly, this coating extends into each well through a distance of approximately one and a half well diameters for contact with the solution and the samples. A heat transfer device is thermally connected to the metallic coating for selectively heating and cooling the samples in the wells of the holding plate.

Abstract: This invention provides methods of obtaining novel polynucleotides and encoded polypeptides by the use of non-stochastic methods of directed evolution (DirectEvolution™). A particular advantage of end-selection-based methods is the ability to recover full-length polynucleotides from a library of progeny molecules generated by mutagenesis methods. These methods include non-stochastic polynucleotide site-saturation mutagenesis (Gene Site Saturation Mutagenesis™) and non-stochastic polynucleotide reassembly (GeneReassembly™). This invention provides methods of obtaining novel enzymes that have optimized physical &/or biological properties. Through use of the claimed methods, genetic vaccines, enzymes, small molecules, and other desirable molecules can be evolved towards desirable properties. For example, vaccine vectors, can be obtained that exhibit increased efficacy for use as genetic vaccines.

Abstract: A purified recombinant phytase enzyme derived from Escherichia coli B. The enzyme has a molecular light of about 47.1 kilodaltons and has phytase activity. The enzyme can be produced from native or recombinant host cells and can be used to aid in the digestion of phytate where desired. In particular, the phytase of the present invention can be used in foodstuffs to improve the feeding value of phytate rich ingredients.

Abstract: Thermostable enzymes are subjected to mutagenesis to produce a thermophilic enzyme which is stable at thermophilic temperature and which has increased activities at least two-fold higher than the activity of the wild-type thermostable enzyme at lower temperatures, which are temperatures of 50° C. or lower.

Abstract: This invention provides methods of obtaining novel polynucleotides and encoded polypeptides by the use of non-stochastic methods of directed evolution (DirectEvolution™). A particular advantage of end-selection-based methods is the ability to recover full-length polynucleotides from a library of progeny molecules generated by mutagenesis methods. These methods include non-stochastic polynucleotide site-saturation mutagenesis (Gene Site Saturation Mutagenesis™) and non-stochastic polynucleotide reassembly (GeneReassembly™). This invention provides methods of obtaining novel enzymes that have optimized physical &/or biological properties. Through use of the claimed methods, genetic vaccines, enzymes, small molecules, and other desirable molecules can be evolved towards desirable properties. For example, vaccine vectors can be obtained that exhibit increased efficacy for use as genetic vaccines.

Abstract: This invention provides methods of obtaining novel polynucleotides and encoded polypeptides by use of non-stochastic methods of directed evolution (DirectEvolution™). These methods include non-stochastic polynucleotide site-saturation mutagenesis (Gene Site Saturation Mutagenesis™) and non-stochastic polynucleotide reassembly (GeneReassembly™). Through use of the claimed methods, genetic vaccines, enzymes, and other desirable molecules can be evolved towards desirable properties. For example, vaccine vectors can be obtained that exhibit increased efficacy for use as genetic vaccines. Vectors obtained by using the methods can have, for example, enhanced antigen expression, increased uptake into a cell, increased stability in a cell, ability to tailor an immune response, and the like. This invention provides methods of obtaining novel enzymes that have optimized physical &/or biological properties.

Abstract: A directed evolution process comprising novel methods for generating improved progeny molecules having desirable properties, including, for example, a method for rapid and facilitated production from a parental polynucleotide template, of a set of mutagenized progeny polynucleotides wherein at least one codon encoding each of the 20 naturally encoded amino acids is represented at each original codon position. This method, termed site-saturation mutagenesis, or simply saturation mutagenesis, is preferably based on the use of the degenerate N,N,G/T sequence. Also, a method of producing from a parental polypeptide template, a set of mutagenized progeny polypeptides wherein each of the 20 naturally encoded amino acids is represented at each original amino acid position.

Abstract: The invention provides novel processes for making ethyl-4-cyano-3-hydroxybutyrate, e.g., (R)-ethyl 4-cyano-3-hydroxybutyric acid, and 4-cyano-3-hydroxybutyric acid. The invention provides protocols for making and 4-cyano-3-hydroxybutyric acid and ethyl-4-cyano-3-hydroxybutyrate by whole cell processes, cell lysate processes, “one pot processes” and “multi-pot” processes using a variety of parameters.

Abstract: In one aspect, invention relates to the field of monitoring chemical reactions and, in one aspect, provides methods and kits for detecting the presence of nitrites and/or cyanide. In one aspect, the invention provides methods and kits for assaying catalytic activity of a material, such as an enzyme, in a chemical or biochemical process, or assaying catalytic activity of a material in a biological sample, such as a whole cell.

Abstract: A directed evolution process comprising novel methods for generating improved progeny molecules having desirable properties, including, for example, a method for rapid and facilitated production from a parental polynucleotide template, of a set of mutagenized progeny polynucleotides wherein at least one codon encoding each of the 20 naturally encoded amino acids is represented at each original codon position. This method, termed site-saturation mutagenesis, or simply saturation mutagenesis, is preferably based on the use of the degenerate N,N,G/T sequence. Also, a method of producing from a parental polypeptide template, a set of mutagenized progeny polypeptides wherein each of the 20 naturally encoded amino acids is represented at each original amino acid position.

Abstract: The invention provides methods for whole cell engineering of new and modified phenotypes by using “on-line” or “real-time” metabolic flux analysis. The invention provides a method for whole cell engineering of new or modified phenotypes by using real-time metabolic flux analysis by making a modified cell by modifying the genetic composition of a cell and culturing the modified cell to generate a plurality of modified cells and measuring at least one metabolic parameter of the cell by monitoring the cell culture of in real time. The invention also provides articles comprising machine-readable medium including machine-executable instructions and systems, e.g., computer systems, to practice the methods of the invention.

Abstract: This invention provides amidases, polynucleotides encoding the amidases, methods of making and using these polynucleotides and polypeptides. In one aspect, the invention provides enzymes having secondary amidase activity, e.g., having activity in the hydrolysis of amides, including enzymes having peptidase, protease and/or hydantoinase activity. In alternative aspects, the enzymes of the invention can be used to used to increase flavor in food (e.g., enzyme ripened cheese), promote bacterial and fungal killing, modify and de-protect fine chemical intermediates, synthesize peptide bonds, carry out chiral resolutions, hydrolyze Cephalosporin C. The enzymes of the invention can be used to generate 7-aminocephalosporanic acid (7-ACA) and semi-synthetic cephalosporin antibiotics, including caphalothin, cephaloridine and cefuroxime. The enzymes of the invention can be used as antimicrobial agents, e.g., as cell wall hydrolytic agents.