酵母双杂交随机肽库的设计及构建

Design and Construction of a Random Peptide Library with Yeast Two-Hybrid System

构建一个含16个氨基酸的酵母双杂交随机肽库,设计合成编码16肽的随机DNA片段,PCR扩增随机DNA片段,经限制性内切酶BamHI和EcoRI酶切后克隆入酵母表达质粒pGADT7GH,构建酵母双杂交随机肽库质粒pGADT7GH-RP并检验其库容。结果表明,扩增获得编码16肽的随机DNA片段,并成功将随机DNA片段克隆入酵母表达质粒pGADT7GH,与GAL4AD形成融合蛋白,其库容为1.28×107。从而成功地构建了酵母双杂交随机肽库。

To construct a peptide library composed of 16 random amino acids with yeast twohybrid system,random oligonucleotides encoded with 16 peptides were designed and artificially synthesized,and random oligonucleotides were amplified by PCR. The amplified products after being digested with BamH I and EcoR I were checked into plasmid pGADT7GH to construct the library plasmids pGADT7GHRP.Then the number of different recombinants were identified.The random oligonucleotides encoded with 16 peptides were obtained and inserted into pGADT7GH,and became GAL4 ADRP fusion protein.The random peptide library with 1.28×107 different recombinant clones was successfully constructed.