口蹄疫病毒3AB基因的克隆与原核表达

Cloning and prokaryotic expression of 3AB gene of FMDV

根据口蹄疫流行毒株设计一对包含3AB部分基因编码区的特异性引物,扩增出3AB基因550 bp的特异带,并将纯化的扩增产物克隆到pM D 18-T载体上。再用设计的酶切位点将3AB基因分别连接到两种原核表达载体上,构建了重组质粒pETCK S-3AB及pET 28a-3AB,转入BL 21菌中进行原核表达。SDS-PAGE电泳表明,3AB基因在两种表达系统中均高效表达。对表达的蛋白通过包涵体及切胶纯化的方法进行纯化,获得了高纯度的3AB蛋白。

According to the polymerase gene of FMDV, the primers of polymerase gene were designed, and the objective fragment was 550 bp which was cloned into pMD18-T vector. Then the polymerase gene was respectively cloned into two prokaryotic expression vectors, and two recombinant plasmids of pETCKS-3AB and pET28a-3AB were constructed. The plasmids were transformed to BL21 （DE3） competent cell, and the polymerase was high--efficiently expressed, The two proteins were purified by the washing of inclusion bodies and gel extraction.