Abstract: Provided are mutant DNA polymerase having at least one mutation which exhibit substantially reduced polymerase activity at 25° C. When compared to the same DNA polymerase without the at least one mutation and which exhibit normal or near-normal polymerase activity at optimum temperatures when compared to the same DNA polymerase without the at least one mutation. Also provided are amino acid sequences and nucleic acid sequences encoding such DNA polymerase, and vector plasmids and host cells suitable for the expression of these sequences. Also described herein are improved methods for performing polymerase chain reaction (PCR) amplification and other genetic manipulations and analyses using the mutant DNA polymerase of the invention.

Abstract: Provided are mutant DNA polymerases having at least one mutation which exhibit substantially reduced polymerase activity at 25° C. when compared to the same DNA polymerases without the at least one mutation and which exhibit normal or near-normal polymerase activity at optimum temperatures when compared to the same DNA polymerases without the at least one mutation. Also provided are amino acid sequences and nucleic acid sequences encoding such DNA polymerases, and vector plasmids and host cells suitable for the expression of these sequences. Also described herein are improved methods for performing polymerase chain reaction (PCR) amplification and other genetic manipulations and analyses using the mutant DNA polymerases of the invention.

Abstract: A method of inhibiting pyrophosphorolysis during DNA chain length elongation is provided. The method comprises including both Mn.sup.++ and Mg.sup.++ in chain-extension and chain-termination reaction mixtures such as those used in DNA sequencing. This method is useful in stabilizing dideoxy-ribonucleoside triphosphate-terminated DNA chains and improving the quality of DNA sequence data obtained via the use of DNA polymerases that do not discriminate against the incorporation of dideoxyribonucleoside triphosphates and other chain terminating agents. Also provided are a reaction mixture and kit for DNA sequencing employing this method.

Abstract: A vector which includes nucleic acid which encodes a DNA polymerase having an identical amino acid sequence to that of the DNA polymerase of Thermus aquaticus termed Taq DNA polymerase, except that it lacks the N-terminal 235 amino acids of Taq DNA polymerase.

Abstract: A DNA polymerase having an amino acid sequence comprising substantially the same amino acid sequence as that of Thermus aquaticus or Thermus flavus DNA polymerase, excluding the N-terminal 280 amino acid residues of Thermus aquaticus DNA polymerase or the N-terminal 279 amino acid residues of Thermus flavus DNA polymerase, recombinant DNA sequences encoding said DNA polymerases, vectors comprising said DNA sequences, and host cells containing such vectors. A formulation of thermostable DNA polymerases comprising a majority component comprised of at least one thermostable DNA polymerase of the type described above, wherein the DNA polymerase lacks 3'-exonuclease activity, and a minority component comprised of at least one thermostable DNA polymerase exhibiting 3'-exonuclease activity, and an improved method for amplifying nucleic acid sequences by polymerase chain reaction wherein the above formulation is mixed and used to catalyze primer extension, are also provided.

Abstract: A support frame with multiple water jets, together with a support grid and carrier frame, is described. Methods for their construction from commonly available components, and methods for their use in a laboratory setting, are provided. This device semi-automates, and improves the quality and reliability of, an otherwise difficult and stressful step in DNA sequencing technology, namely the transfer of a thin and fragile gel from a glass plate to a sheet of filter paper for subsequent drying under vacuum.