Compositions and methods utilizing DNA polymerases
AssigneeAgilent Technologies, Inc.
InventorsJoseph A. Sorge, Connie Jo Hansen, Holly Hogrefe
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Abstract
The invention features a novel isolated Family B DNA polymerase, a Thermococcus polymerase JDF-3, and mutant recombinant forms thereof. Mutant polymerases of the invention are deficient in 3′ to 5′ exonuclease activity and/or exhibit reduced discrimination against non-conventional nucleotides relative to the wild-type form of the polymerase.
Claims66
A composition for identifying a nucleotide at a given position of a template DNA molecule, said composition comprising a Family B DNA polymerase having reduced discrimination against non-conventional nucleotides and a first primer, wherein said first primer anneals to the immediate 3′ of said nucleotide at the given position of said template DNA molecule, and wherein said Family B DNA polymerase has a substitution at a position corresponding to L408, P410, or both L408 and P410 of SEQ ID NO:2, which result in reduced discrimination by the mutant polymerase as compared to the non-mutant polymerase from which it is derived.
An isolated recombinant Family B DNA polymerase having reduced discrimination against non-conventional nucleotides, wherein said DNA polymerase comprises an amino acid substitution at a position corresponding to P410 of SEQ ID NO:2, which reduces discrimination by the mutant polymerase as compared to the non-mutant polymerase from which it is derived.
A kit for identifying a nucleotide at a given position of a template DNA molecule, said kit comprising a Family B DNA polymerase having reduced discrimination against non-conventional nucleotides and a first primer, wherein said first primer anneals to the immediate 3′ of said nucleotide at the given position of said template DNA molecule, and wherein said Family B DNA polymerase has a substitution at a position corresponding to L408, P410, or both L408 and P410 of SEQ ID NO:2, which result in reduced discrimination by the mutant polymerase as compared to the non-mutant polymerase from which it is derived.
A kit for identifying a nucleotide at a given position of a template DNA molecule, said kit comprising a Family B DNA polymerase having reduced discrimination against non-conventional nucleotides, wherein said DNA polymerase comprises an amino acid substitution at a position corresponding to P410 of SEQ ID NO:2, which results in reduced discrimination by the mutant polymerase as compared to the non-mutant polymerase from which it is derived.
Dependencies are inferred from the claim text.