Mutant <i>Neq </i>HS DNA polymerase derived from <i>Nanoarchaeum equitans </i>and its application to hot-start PCR

摘要

A DNA polymerase (Neq DNA polymerase) derived from Nanoarchaeum equitans is split into Neq L and Neq S fragments, each of which contains inteins. A Neq hot-start (HS) DNA polymerase in which the inteins of the Neq L and Neq S fragments are linked with each other is provided in the form of a precursor of Neq DNA polymerase. A purification method can be significantly improved by inserting a His-tag sequence composed of six histidine residues between the inteins of the Neq L and Neq S fragments at a gene level. As a result of effort to enhance PCR efficiency of the Neq HS DNA polymerase, a gene coding for the Neq HS DNA polymerase is mutated at specific positions to screen mutant Neq HS polymerases (M1, M2, and M3) having a highly improved PCR amplification rate and amplification level.

主权项

1. A thermostable hot-start DNA polymerase, wherein the thermostable hot-start DNA polymerase comprises the amino acid sequence set forth in SEQ ID NO: 6, SEQ ID NO: 32, SEQ ID NO: 34, or SEQ ID NO: 36.