H5N1型流感病毒M2蛋白全长编码基因的克隆与原核表达

Clone and expression of M2 protein of influenza A H5N1 virus

目的:构建H5N1流感病毒基质蛋白M2基因的原核表达系统,为研究其在体液免疫中的作用奠定物质基础。方法:应用PCR方法从流感病毒H5N1cDNA中扩增出M2的全基因序列,插入至表达质粒载体pGEX6P-3上,构建重组原核表达质粒pGEX-6p-3/M2。重组质粒经测序分析确认后,转化感受态大肠杆菌BL21,经IPTG诱导表达M2蛋白后,对其表达产物进行SDS-PAGE和Western Bloting分析。结果:PCR扩增的M2基因长度为297bp,经酶切和测序分析,插入到载体的基因与GeneBank公布的序列相似性达100%;SDS-PAGE和Westem Bloting分析显示,经IPTG诱导出分子质量为37.2 KD的目的蛋白。结论:成功构建了H5N1流感病毒基质蛋白M2原核表达载体pGEX-6p-3/M2,并在大肠杆菌中获得表达。

Objective：To clone and express the gene of matrix protein 2（M2） of H5N1 influenza virus,and to establish material foundation for the study of function mechanism in body fluid immunity.Methods：The M2 gene whose length was 297 bp was amplified by PCR from cDNA of influenza virus,and cloned into the expression vector pGEX-6P-3.Recombinant plasmid was then transformed into E.coli BL21,which was induced with IPTG.The whole protein of E.coli was analyzed by SDS-PAGE and western bloting.Results：M2 protein of influenza A virus,whose weight was 37.2 kD was successfully cloned and expressed in the E.coli.Conclusion：M2 protein acquired good fusion expression,which can provide basis for the further study on its immunity mechanism.