Helicase dependent isothermal amplification using nicking enzymes

摘要

The present invention relates to a method for amplifying a template nucleic acid, wherein the method comprises amplifying said template nucleic acid using the helicase dependent amplification (HDA) reaction in the presence of a nicking endonuclease, and wherein said template nucleic acid comprises a sequence recognized by said nicking endonuclease or a sequence recognized by said nicking endonuclease is introduced into the template nucleic acid during the HDA reaction. The invention further pertains to a kit for amplifying a nucleic acid, comprising a nicking endonuclease, a helicase and a DNA polymerase.

主权项

1. A method for amplifying a template nucleic acid, comprising:
(a) providing a template nucleic acid, (b) contacting the template nucleic acid with a helicase, (c) hybridizing an oligonucleotide primer to the template nucleic acid, and (d) amplifying said template nucleic acid using the helicase dependent amplification (HDA) reaction, said HDA reaction introducing a recognition sequence into the template nucleic acid, said HDA reaction occurring in the presence of a nicking endonuclease that recognizes a nucleotide sequence of a completely or partially double-stranded nucleic acid molecule and cleaves only one strand of the template nucleic acid at a specific location relative to the recognition sequence, wherein the HDA reaction is a thermophilic HDA (tHDA), and wherein the template nucleic acid is shorter than 400 bp.