Abstract: The present invention relates to a DNA and RNA polymerases which have increased fidelity (or reduced misincorporation rate). In particular, the invention relates to a method of making such polymerases by modifying or mutating the nucleotide binding domain of the polymerase (e.g., the O-helix). The invention also relates to DNA molecules containing the genes encoding the polymerases of the invention, to host cells containing such DNA molecules and to methods to make the polymerases using the host cells. The polymerases are particularly suited for nucleic acid synthesis, sequencing, amplification and cDNA synthesis.

Abstract: The present invention is directed generally to methods facilitating the cloning of nucleic acid molecules. In particular, the invention relates to the use of polymerase inhibitors, including but not limited to anti-polymerase antibodies (such as anti-Taq antibodies) and fragments thereof, to inactivate residual polymerase activity remaining after the amplification (particularly via PCR) of a target nucleic acid molecule. The invention further provides compositions, particularly storage-stable compositions, comprising one or more components, such as one or more restriction endonucleases and one or more polymerase inhibitors, that are useful in cloning amplified or synthesized nucleic acid molecules by the above-described methods. The invention also relates to nucleic acid molecules produced by these methods, and to genetic constructs (such as vectors) and host cells comprising these nucleic acid molecules.

Abstract: The present invention relates to nucleic acid inhibitors, compositions and method for enhancing synthesis of nucleic acid molecules. In a preferred aspect, the invention relates to inhibition or control of nucleic acid synthesis, sequencing or amplification. Specifically, the present invention discloses nucleic acids having affinity for polypeptides with polymerase activity for use in such synthesis, amplification or sequencing reactions. The nucleic acid inhibitors are capable of inhibiting nonspecific nucleic acid synthesis under certain conditions (e.g., at ambient temperatures). Thus, in a preferred aspect, the invention relates to “hot start” synthesis of nucleic acid molecules. Accordingly, the invention prevents, reduces or substantially reduces nonspecific nucleic acid synthesis. The invention also relates to kits for synthesizing, amplifying, reverse transcribing or sequencing nucleic acid molecules comprising one or more of the nucleic acid inhibitors or compositions of the invention.

Abstract: The present invention relates to mutant DNA polymerases which incorporate dideoxynucleotides with about the same efficiency as deoxynucleotides. The present invention also related to mutant DNA polymerases which also have substantially reduced 5?-to-3? exonuclease activity or 3?-to-5? exonuclease activity. The invention also relates to DNA molecules coding for the mutant DNA polymerases, and hosts containing the DNA molecules.

Abstract: The invention relates to a substantially pure thermostable DNA polymerase from Thermotoga (Tne and Tma) and mutants thereof. The Tne DNA polymerase has a molecular weight of about 100 kilodaltons and is more thermostable than Taq DNA polymerase. The mutant DNA polymerase has at least one mutation selected from the group consisting of (1) a first mutation that substantially reduces or eliminates 3??5? exonuclease activity of said DNA polymerase; (2) a second mutation that substantially reduces or eliminates 5??3? exonuclease activity of said DNA polymerase; (3) a third mutation in the O helix of said DNA polymerase resulting in said DNA polymerase becoming non-discriminating against dideoxynucleotides. The present invention also relates to the cloning and expression of the wild type or mutant DNA polymerases in E. coli, to DNA molecules containing the cloned gene, and to host cells which express said genes.

Abstract: The present invention provides methods for use in identifying, analyzing and typing polymorphic DNA fragments, particularly minisatellite, microsatellite or STR DNA fragments. In particular, the invention provides methods using DNA polymerases, more particularly thermostable DNA polymerases, and most particularly Thermotoga polymerases or mutants or derivatives thereof, whereby minisatellite, microsatellite or STR DNA molecules may be amplified and analyzed for polymorphisms. The invention also relates to polymerases having reduced, substantially reduced or eliminated ability to add non-template 3? nucleotides to a synthesized nucleic acid molecule. In accordance with the invention, such reduction or elimination may be accomplished by modifying or mutating the desired polymerase.

Abstract: The invention provides a microarray and methods for producing a protein microarray. The array comprises multiple nucleic acid molecules immobilized on a substrate, each comprising (i) a protein-binding domain and (ii) a nucleic acid sequence encoding a fusion protein comprising a polypeptide of interest and a DNA-binding protein that binds the protein-binding domain, and one or more fusion proteins produced from the multiple nucleic acid molecules. Each fusion protein is immobilized on the substrate via binding to a nucleic acid sequence comprising the protein-binding domain present on the nucleic acid molecule from which the fusion protein is produced or on the substrate. The invention also provides a method of analyzing protein interactions with, for example, other proteins, lipids and drugs.

Abstract: The present invention relates to nucleic acid inhibitors, compositions and method for enhancing synthesis of nucleic acid molecules. In a preferred aspect, the invention relates to inhibition or control of nucleic acid synthesis, sequencing or amplification. Specifically, the present invention discloses nucleic acids having affinity for polypeptides with polymerase activity for use in such synthesis, amplification or sequencing reactions. The nucleic acid inhibitors are capable of inhibiting nonspecific nucleic acid synthesis under certain conditions (e.g., at ambient temperatures). Thus, in a preferred aspect, the invention relates to “hot start” synthesis of nucleic acid molecules. Accordingly, the invention prevents, reduces or substantially reduces nonspecific nucleic acid synthesis. The invention also relates to kits for synthesizing, amplifying, reverse transcribing or sequencing nucleic acid molecules comprising one or more of the nucleic acid inhibitors or compositions of the invention.

Abstract: The current invention provides methods for producing a polypeptide as inclusion bodies in bacterial host cells. The present methods are carried out by forming a gene construct comprising the genetic sequence encoding a polypeptide operatively linked to that of an inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such that host cells comprising the gene construct produce the polypeptide as intracellular inclusion bodies. The methods of the present invention facilitate the rapid isolation and purification of recombinant proteins. In addition, the present methods may be useful for producing polypeptides or proteins which are small and are typically difficult to express, as well as those proteins that are toxic to host cells such as E. coli. The present invention also provides plasmids, vectors and host cells to be used in the present invention for production of polypeptides, and methods of production of polypeptides using these vectors and host cells.

Abstract: The current invention provides methods for producing a polypeptide as inclusion bodies in bacterial host cells. The present methods are carried out by forming a gene construct comprising the genetic sequence encoding a polypeptide operatively linked to that of an inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such that host cells comprising the gene construct produce the polypeptide as intracellular inclusion bodies. The methods of the present invention facilitate the rapid isolation and purification of recombinant proteins. In addition, the present methods may be useful for producing polypeptides or proteins which are small and are typically difficult to express, as well as those proteins that are toxic to host cells such as E. coli. The present invention also provides plasmids, vectors and host cells to be used in the present invention for production of polypeptides, and methods of production of polypeptides using these vectors and host cells.

Abstract: The current invention provides methods for producing a polypeptide as inclusion bodies in bacterial host cells. The present methods are carried out by forming a gene construct comprising the genetic sequence encoding a polypeptide operatively linked to that of an inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such that host cells comprising the gene construct produce the polypeptide as intracellular inclusion bodies. The methods of the present invention facilitate the rapid isolation and purification of recombinant proteins. In addition, the present methods may be useful for producing polypeptides or proteins which are small and are typically difficult to express, as well as those proteins that are toxic to host cells such as E. coli. The present invention also provides plasmids, vectors and host cells to be used in the present invention for production of polypeptides, and methods of production of polypeptides using these vectors and host cells.

Abstract: A method of producing a Rous Sarcoma Virus reverse transcriptase (RSV RT) by expressing one orniore nucleic acid sequences encoding one or more subunits of RSV RT in a eukaryotic host cell and culturing the host cell under conditions sufficient to produce the recombinant RSV RT. The resulting RSV RT has a specific activity of between about 30,000 and 150,000 units per milligram and is suitable for methods including RT-polyinerase chain reaction (RT-PCR).

Abstract: The invention relates to compositions comprising mixtures of polypeptides having reverse transcriptase (RT) activity and to methods of producing, amplifying or sequencing nucleic acid molecules using these compositions or polypeptides, particularly at temperatures above about 55° C. The invention also relates to nucleic acid molecules produced by these methods, to vectors and host cells comprising these nucleic acid molecules, and use of such nucleic acid molecules to produce desired polypeptide. The invention also relates to methods for producing Avian Sarcoma-Leukosis Virus (ASLV) RT subunits, in particular, Avian Myeloblastosis Virus (AMV) RTs, to isolated nucleic acid molecules encoding ASLV RT subunits, and to ASLV RT subunits produced by these methods. The invention further relates to nucleic acid molecules encoding recombinant RT holoenzymes, particularly ASLV RTs, methods for producing these RTs and to RTs produced by these methods.

Abstract: The invention relates to compositions comprising mixtures of polypeptides having reverse transcriptase (RT) activity and to methods of producing, amplifying or sequencing nucleic acid molecules using these compositions or polypeptides, particularly at temperatures above about 55° C. The invention also relates to nucleic acid molecules produced by these methods, to vectors and host cells comprising these nucleic acid molecules, and use of such nucleic acid molecules to produce desired polypeptide. The invention also relates to methods for producing Avian Sarcoma-Leukosis Virus (ASLV) RT subunits, in particular, Avian Myeloblastosis Virus (AMV) RTs, to isolated nucleic acid molecules encoding ASLV RT subunits, and to ASLV RT subunits produced by these methods. The invention further relates to nucleic acid molecules encoding recombinant RT holoenzymes, particularly ASLV RTs, methods for producing these RTs and to RTs produced by these methods.

Abstract: The present invention relates to nucleic acid inhibitors, compositions and method for enhancing synthesis of nucleic acid molecules. In a preferred aspect, the invention relates to inhibition or control of nucleic acid synthesis, sequencing or amplification. Specifically, the present invention discloses nucleic acids having affinity for polypeptides with polymerase activity for use in such synthesis, amplification or sequencing reactions. The nucleic acid inhibitors are capable of inhibiting nonspecific nucleic acid synthesis under certain conditions (e.g., at ambient temperatures). Thus, in a preferred aspect, the invention relates to “hot start” synthesis of nucleic acid molecules. Accordingly, the invention prevents, reduces or substantially reduces nonspecific nucleic acid synthesis. The invention also relates to kits for synthesizing, amplifying, reverse transcribing or sequencing nucleic acid molecules comprising one or more of the nucleic acid inhibitors or compositions of the invention.

Abstract: The current invention provides methods for producing a polypeptide as inclusion bodies in bacterial host cells. The present methods are carried out by forming a gene construct comprising the genetic sequence encoding a polypeptide operatively linked to that of an inclusion partner protein, such as E. coli thioredoxin or a modified E. coli thioredoxin, such that host cells comprising the gene construct produce the polypeptide as intracellular inclusion bodies. The methods of the present invention facilitate the rapid isolation and purification of recombinant proteins. In addition, the present methods may be useful for producing polypeptides or proteins which are small and are typically difficult to express, as well as those proteins that are toxic to host cells such as E. coli. The present invention also provides plasmids, vectors and host cells to be used in the present invention for production of polypeptides, and methods of production of polypeptides using these vectors and host cells.

Abstract: The present invention is generally related to compositions and methods for the reverse transcription of nucleic acid molecules, especially messenger RNA molecules. Specifically, the invention relates to compositions comprising mixtures of polypeptides having reverse transcriptase (RT) activity, and to methods of producing, amplifying or sequencing nucleic acid molecules (particularly cDNA molecules) using these compositions or polypeptides, particularly at temperatures above about 55° C. The invention also relates to nucleic acid molecules produced by these methods, to vectors and host cells comprising these nucleic acid molecules, and to the use of such nucleic acid molecules to produce desired polypeptides.

Abstract: The present invention is generally related to compositions and methods for the reverse transcription of nucleic acid molecules, especially messenger RNA molecules. Specifically, the invention relates to compositions comprising mixtures of polypeptides having reverse transcriptase (RT) activity, and to methods of producing, amplifying or sequencing nucleic acid molecules (particularly cDNA molecules) using these compositions or polypeptides, particularly at temperatures above about 55° C. The invention also relates to nucleic acid molecules produced by these methods, to vectors and host cells comprising these nucleic acid molecules, and to the use of such nucleic acid molecules to produce desired polypeptides.

Abstract: The invention relates to a substantially pure thermostable DNA polymerase from Thermotoga (Tne and Tma) and mutants thereof. The Tne DNA polymerase has a molecular weight of about 100 kilodaltons and is more thermostable than Taq DNA polymerase. The mutant DNA polymerase has at least one mutation selected from the group consisting of (1) a first mutation that substantially reduces or eliminates 3′→5′ exonuclease activity of said DNA polymerase; (2) a second mutation that substantially reduces or eliminates 5′→3′ exonuclease activity of said DNA polymerase; (3) a third mutation in the O helix of said DNA polymerase resulting in said DNA polymerase becoming non-discriminating against dideoxynucleotides. The present invention also relates to the cloning and expression of the wild type or mutant DNA polymerases in E. coli, to DNA molecules containing the cloned gene, and to host cells which express said genes.

Abstract: The invention relates to a substantially pure thermostable DNA polymerase from Thermotoga neapolitana (Tne). The Tne DNA polymerase has a molecular weight of about 100 kilodaltons and is more thermostable than Taq DNA polymerase. The present invention also relates to the cloning and expression of the Tne DNA polymerase in E. coli, to DNA molecules containing the cloned gene, and to host cells which express said genes. The Tne DNA polymerase of the invention may be used in well-known DNA sequencing and amplification reactions.