CPC C12N 9/1252 (2013.01) [C12P 19/34 (2013.01); C12Q 1/6869 (2013.01); C12Y 207/07007 (2013.01)] | 20 Claims |
1. A method for incorporating a single nucleotide into a priming strand in a template-independent reaction, the method comprising:
combining a priming strand with a 3′-OH-unmodified reversible terminator and a mutant polymerase, wherein the mutant polymerase is at least 96% identical to SEQ ID NO:2 and comprises:
a Y546H mutation at a position functionally equivalent to position 546 in Pfu polymerase;
a L409Y, L409H or L409F mutation at a position functionally equivalent to position 409 in Pfu polymerase; and
a A486X mutation at a position functionally equivalent to position 486 in Pfu polymerase, wherein X is any amino acid except alanine;
wherein incorporation of the terminator is at least 2-fold higher than for the mutant DNA polymerase of SEQ ID NO:11.
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