Patent ID: 8580507
Filing Date: 2013-11-12
Classification: C12N,C12Q,G01N

Abstract:
1. A process comprising: (a) contacting a recombinase agent with a first and a second nucleic acid primer and a third extension blocked primer which comprises a noncomplementary or modified internal residue to form a first, second and third nucleoprotein primer; (b) contacting the first and second nucleoprotein primers to a double stranded target nucleic acid comprising a first and second strand, thereby forming a first double stranded structure between said first nucleoprotein primer and said first strand at a first portion of said first strand and a second double stranded structure between said second nucleoprotein primer and said second strand at a second portion of said second strand such that the 3′ ends of said first nucleoprotein primer and said second nucleoprotein primer are oriented toward each other on the same target nucleic acid with a third portion of target nucleic acid between said 3′ ends; (c) extending the 3′ end of said first nucleoprotein primer and second nucleoprotein primer with one or more polymerases and dNTPs to generate an amplified target nucleic acid with an internal region comprising the third portion of nucleic acid; and (d) contacting said amplified target nucleic acid to said third nucleoprotein primer to form a third double stranded structure at the third portion of said amplified target nucleic acid in the presence of a nuclease; wherein said nuclease specifically cleaves said noncomplementary or modified internal residue only after the formation of said third double stranded structure to form a third 5′ primer double stranded structure and a third 3′ extension blocked primer double stranded structure.