摘要
实验以汉坦病毒76-118株M基因为模板进行PCR扩增得到含完整编码汉坦病毒G2蛋白基因M2的1 600 bp DNA片段,将此基因片段重组到T载体pMD18中,转化至大肠埃希菌DH5α后,筛选阳性菌落扩增培养,提取重组质粒,双酶切鉴定后,克隆至表达载体pGEX-6P-1-M2,亲和层析法纯化融合蛋白,产物进行Western-Blot鉴定。实验证实通过以质粒pGEX-6P-1为载体,可构建编码汉坦病毒76-118株包膜糖蛋白G2的M2基因的原核表达载体pGEX-6P-1-M2,并可观察G2蛋白在原核细胞中的表达。
In this experiment, Hantaan virus 76-118 strain M gene was served as a template for PCR amplification to get 1 600 bp DNA fragments of Hantaan virus G2 protein gene containing integral coding information. The fragments were recombined into T vector of pMD18 and then after transformed into E. coil DHSα to screen positive colonies and amplify and culture to extract the recombinant plasmid, which was identified by double digestion method. The product was then sub-cloned into pGEX-6P-1-M2 expression vector. The fusion protein was then purified by affinity chromatog- raphy, finally the protein was identified by Western-Blot. This experiment had proved that taking plasmid pGEX-6P-1 as a vector could construct a prokaryotic expression vector pGEX-6P-1-M2 eneoding Hantaan virus 76-118 strain M gene of involucral glycoprotein G2, and could observe G2 protein expression in prokaryotic cells.
出处
《微生物学杂志》
CAS
CSCD
2012年第6期29-32,共4页
Journal of Microbiology
基金
辽宁省教育厅2012年科学研究一般项目(L2012287)
关键词
汉坦病毒
G2蛋白
原核表达载体
蛋白表达
Hantaan virus
G2 protein
prokaryotic expression vector
protein expression
