论文部分内容阅读
目的克隆并构建耐甲氧西林金黄色葡萄球菌(MRSA)青霉素结合蛋白2a(PBP2a)全长及转肽酶区的原核表达质粒。方法登录基因文库查找获得mecA基因的编码序列,应用PCR技术扩增获得DNA片段,将此基因片段插入PET-32a载体,同时酶切鉴定阳性克隆,DNA序列测定验证序列正确性。结果 PCR扩增获得了mecA基因全长及转肽酶区DNA片段,成功插入到原核表达载体PET32a,双酶切鉴定及DNA序列测定证实插入片段正确。结论成功构建了PBP2a全长及转肽酶区片段表达质粒,为该蛋白的纯化表达和疫苗研究奠定了基础。
Objective To clone and construct a full-length prokaryotic expression plasmid of methicillin-resistant Staphylococcus aureus (MRSA) penicillin binding protein 2a (PBP2a) and transpeptidase. Methods The coding sequence of mecA gene was obtained by searching the gene library. The DNA fragment was amplified by PCR. The gene fragment was inserted into PET-32a vector and positive clones were identified by restriction enzyme digestion. DNA sequencing confirmed the correctness of the sequence. Results The full length of mecA gene and DNA fragment of transpeptidase were obtained by PCR. The recombinant plasmid was successfully inserted into prokaryotic expression vector PET32a. Double enzyme digestion and DNA sequencing confirmed that the inserted fragment was correct. Conclusion The full-length PBP2a fragment and transpeptidase region expression plasmid was successfully constructed, which laid the foundation for the purification of the protein and vaccine research.