生殖支原体MG 427原核表达载体的构建和鉴定

目的构建生殖支原体MG 427基因原核表达载体并进行鉴定。方法以生殖支原体标准株G 37 DNA为模板进行PCR扩增mg 427基因,克隆入原核表达载体p GEX-6 p-1。定点诱导该基因中第289-291位核苷酸TGA密码子突变成TGG,构建p GEX-6 p-1/MG 427的原核表达载体。结果成功扩增出大约426 bp的基因片段,定点诱导突变后,经酶切及测序鉴定证明mg 427中的TGA被成功突变为TGG,与目的基因相符。结论成功构建生殖支原体渗透压诱导蛋白MG 427原核表达载体p GEX-6 p-1/MG 427。

Objective To construct and identified recombinant prokaryotic vector containing the gene MG 427 of Mycoplasma genitalium. Methods mg427 gene was amplified by polymerase chain reaction （PCR） with M, genitalium strain G 37 DNA for the template. The PCR products were directly cloned into pGEX-6 p-1 expression vector to constructed recombinant plasmid pGEX-6 p-1/mg 427. A codon TGA in the position of 289-291 of gene was muted into TGG by site-directed mutagenesis. Results The target gene successfully amplified. The results of PCR and sequencing showed that the amplified DNA sequences were consistent with the sequences of the mg427 gene in GenBank. The TGA in position of 289-291 of mg427 gene was successly mutated into TGG. By PCR-mediated site-directed mutagenesis. Conclusion Prokaryotic expression vector pGEX-6 p-1/mg 427 was constructed successfully.