Balb/C小鼠金属硫蛋白-1基因在大肠杆菌中的高效表达

摘要

将Balb/C小鼠金属硫蛋白-1基因克隆于质粒表达载体pET-21a 上，克隆采用限制性内切酶BamH1与EcoR1酶切pET21a及金属硫蛋白-1基因PCR产物，连接成 pET21a-MT重组质粒，并将其转化进入大肠杆菌表达受体菌BL21(DE3)中，经异丙基-β-D -硫代半乳糖苷(IPTG)诱导，SDS-聚丙烯酰胺凝胶电泳(PAGE)证实产生高效表达金属硫蛋白-1 基因的金属硫蛋白融合蛋白。表达蛋白在胞内主要以不溶性包涵体存在。%Mouse Balb/C metallothionein cDNA was cloned into pET21 a with the rest riction endonuclease BamH1 and EcoR1 to form the recombinant plasmid pET21a-MT . pET21a-MT was then transformed into the BL21(DE3).After the induction of IPTG , the protein produced from the BL21(pET21a-MT) was proved to be the metallothion e in fusion protein (99 amino acids/per protein molecule).SDS-PAGE showed that the induced protein was present mainly as the inclusion body in E.coli.