结核分枝杆菌Rv2389基因真核表达质粒的构建及表达

Construction and expression of eukaryotic expression vector of Mycobacterium tuberculosis Rv2389

目的:构建结核分枝杆菌Rv2389基因真核表达载体.方法:PCR扩增Rv2389基因,测序正确后克隆入真核表达载体pCDNA3.1(-),重组质粒酶切鉴定正确后以阳离子聚合物转染CHO细胞后,分别以RT-PCR方法检测mRNA表达和间接免疫荧光技术检测目的蛋白的表达.结果:构建了重组质粒pCDNA-Rv2389,RT-PCR结果证明Rv2389可在CHO细胞中转录,间接免疫荧光检测证明,表达有Rv2389蛋白的细胞着染.结论:成功构建了结核分枝杆菌Rv2389基因的真核表达载体pCDNA-Rv2389,Rv2389基因可以在CHO细胞中表达.

AIM： To construct the eukaryotic expression vector encoding Mycobacterium tuberculosis Rv2389 and express it in CHO cells. METHODS： The gene encoding Rv2389 protein were amplified by polymerase chain reaction （ PCR ） from genome of Mycobacterium tuberculosis H37Rv strain. After sequenced, Rv2389 gene segments were subcloned into eukaryotic expression vector pCDNA3. 1 （ - ）. The recombinant plasmid pCDNA- Rv2389 were transfected into CHO cells with liposome. The expressions of mRNA and protein encoded by this gene were detected respectively with RT-PCR and indirect immunofluoresent technology. RESULTS： Rv2389 was cloned into pCDNA3. 1 （ - ） correctly, and its expression at mRNA and protein levels was detected in CHO cells. CONCLUSION： Recombinant eukaryotic plasmid encoding Rv2389 was constructed successfully. The experiment established the basis for further study on the function of Rv2389.