Patent ID: 6197556
Filing Date: 2001-03-06
Classification: C12Q

Abstract:
A method for amplifying a segment of a nucleic acid template, said method comprising:(a) annealing the template to a first branched primer, said primer comprising a front and a back oligonucleotide module, wherein(i) the front oligonucleotide nodule comprises a stem segment;(ii) said stem segment is complementary to a stem segment of the back oligonucleotide module,(iii) said front and back oligonucleotide modules anneal to each other by means of said stem segments;(iv) each of said oligonucleotide modules also comprises an arm segment, that is complementary to a site in the template; and(v) said site to which the arm segments of the front and back modules are complementary, and are sufficiently close to each other in the template so that said first branched primer forms a 3-way junction when annealed to the template;(b) extending the arm of the front module of the first branched primer on the temple by a polymerase enzyme to form a first initial extension strand;(c) annealing the first initial extension strand to a reverse primer which is capable of extension in the direction of the 5' end of the first initial extension strand;(d) extending said reverse primer on the first initial extension strand by a polymerase enzyme to form a second initial extension stand, which strand includes a complement to the front module of the first branched primer; and(e) amplifying a segment of the second initial extension strand comprising a copy of the segment of the nucleic acid template wherein the template segment is amplified by using amplification primers.