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AssigneeInvitrogen Corporation
InventorsJoseph Solus, Shuwei Yang, Deb K. Chatterjee
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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 maybe 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.
A polymerase which comprises one or more modifications or mutations in the O-helix of said polymerase that reduce, substantially reduce or eliminate the ability of the polymerase to add non-templated 3′ nucleotides to a synthesized nucleic acid molecule, wherein said polymerase is not a Thermotoga polymerase.
A mutant Tne DNA polymerase protein selected from the group consisting of:
A polymerase which comprises one or more modifications or mutations in the O-helix of said polymerase that reduce, substantially reduce or eliminate the ability of the polymerase to add non-templated 3′ nucleotides to a synthesized nucleic acid molecule, wherein said mutation or modification is at one or more positions or combinations of positions of said O-helix selected from the group consisting of position R (Arg), position F (Phe), position K (Lys), positions R and F, positions R and K, positions F and K, and positions R, F and K, and wherein said polymerase is not a Thermotoga polymerase.
A polymerase which comprises one or more modifications or mutations in the O-helix of said polymerase that reduce, substantially reduce or eliminate the ability of the polymerase to add non-templated 3′ nucleotides to a synthesized nucleic acid molecule, wherein said mutation or modification is an amino acid substitution at one or more positions or combinations of positions of said O-helix selected from the group consisting of position R, position F, position K, positions R and F, positions R and K, positions F and K, and positions R, F and K, and wherein said polymerase is not a Thermotoga polymerase.
Dependencies are inferred from the claim text.