耐甲氧西林金黄色葡萄球菌mecA基因原核表达载体的构建

目的对耐甲氧西林的金黄色葡萄球菌(MRSA)mecA基因片段进行扩增表达,纯化及鉴定。方法应用聚合酶链反应(PCR)扩增技术获得编码PBP2a转肽酶区的mecA基因片段,将此目的基因片段与pET-21a(+)载体连接,构建pET-21a(+)-mecA的重组质粒,经双酶切、测序正确后,将重组质粒转入E.coli BL21 DE3中,用IPTG诱导mecA融合蛋白,并对表达产物进行鉴定。结果构建了mecA基因的原核表达载体,并获得高效表达。结论获得了mecA基因编码的PBP2a蛋白,为MRSA的耐药机制、药物治疗等进一步研究奠定了基础。

Construction of a prokaryotic expression vector of mecA gene in MRSA

Objective To amplifiy, express, puffy and indentify the mecA gene fragment of methicillin-resistant Staphylococcus aureus （MRSA）. Methods PCR amplification was used to obtain mecA gene fragment containing transpeptidase PBP2a coding region, which was conjugated with pET-21a （＋） carrier to establish recombinant plasmid of pET-21a （＋）-mecA. After double enzyme digestion and sequence verification, the recombinant plasmid was trans- formed into E.coli BL21 （DE3）. IPTG induction of mecA fusion protein was performed and expression products were verified. Results The prokaryotic expression vector of mecA gene was established with a highly efficient expression. Conclusion An mecA gene fragment coding PBP2a protein was obtained, which may provide a basis for further study of drug resistance and medical treatment of MRSA.