Modified polymerases for improved incorporation of nucleotide analogues
First Claim
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1. An altered family B archaeal DNA polymerase, wherein the second amino acid of the motif A region is mutated to alanine (A);
- and the third amino acid of the motif A region is mutated to an amino acid selected from the group consisting of serine (S), alanine (A), glycine (G), and the amino acids having beta-branched side chains, whereby the altered family B archaeal DNA polymerase is capable of incorporating a 3′
O-azidomethyl-modified nucleotide.
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Abstract
The invention relates to modified polymerase enzymes which exhibit improved incorporation of nucleotide analogs bearing substituents at the 3′ position of the sugar moiety that are larger in size than the naturally occurring 3′ hydroxyl group. Also described are methods of using the polymerases to incorporate nucleotides into polynucleotides, particularly in the context of DNA sequencing.
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Citations
21 Claims
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1. An altered family B archaeal DNA polymerase, wherein the second amino acid of the motif A region is mutated to alanine (A);
- and the third amino acid of the motif A region is mutated to an amino acid selected from the group consisting of serine (S), alanine (A), glycine (G), and the amino acids having beta-branched side chains, whereby the altered family B archaeal DNA polymerase is capable of incorporating a 3′
O-azidomethyl-modified nucleotide. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21)
- and the third amino acid of the motif A region is mutated to an amino acid selected from the group consisting of serine (S), alanine (A), glycine (G), and the amino acids having beta-branched side chains, whereby the altered family B archaeal DNA polymerase is capable of incorporating a 3′
Specification