公开(公告)号：US20200248231A1

公开(公告)日：2020-08-06

申请号：US16641946

申请日：2018-08-28

Applicant: Spiber Inc.

Inventor： Yusuke Ito ,  Chikako Sato ,  Nozomu Yachie

IPC: C12Q1/686

Abstract: The present invention relates to a method for producing a double-stranded DNA fragment having a desired nucleotide sequence through dual asymmetric PCR (DA-PCR). The method comprises: (1) providing a plurality of oligonucleotides (sense oligonucleotides) each corresponding to a part of a sense strand of the double-stranded DNA fragment and a plurality of oligonucleotides (antisense oligonucleotides) each corresponding to a part of an antisense strand of the double-stranded DNA fragment and mixing together the oligonucleotides with equal concentrations, DNA polymerase, and dNTP to prepare a reaction mixture solution; (2) performing PCR by using the reaction mixture solution from step (1); (3) adding a primer set capable of amplifying the double-stranded DNA fragment of full length to the reaction mixture solution from step (2); and (4) performing PCR by using the reaction mixture solution from step (3).