牛分支杆菌ag85b基因的克隆及表达

Cloning and Prokaryotic Expression of ag85b Gene of Mycobacterium Bovis

以牛分支杆菌C68001株基因组DNA为模板,克隆免疫优势抗原基因ag85b,构建重组表达质粒pET-28a-ag85b,转化入大肠埃希菌BL21(DE3),IPTG诱导表达重组蛋白,对表达产物进行SDS-PAGE分析和Western blot鉴定。结果表明,构建了ag85b基因的重组基因工程菌株,IPTG诱导表达的重组蛋白分子质量约为33ku,目的蛋白能被His单克隆抗体识别,表达产物主要以包涵体形式存在。

To clone and express the geneen coding Mycobacterium tuberculosis antigen Ag85B, a pair of spe- cific primers were designed, the Ag85B gene was amplified from the genomic DNA by PCR, and cloned in- to vector PET28a （＋） to construct the recombinant plasmid. The recombinant plasmid was transformed into the expression vector Escherichia coli BL21 （DE3）, and induced with IPTG; the presence of the recombinant protein in the expression vector was analysed by SDS-PAGE and Western blot. Results showed that the ag85b gene was successfully expressed with expected 33 ku in molecular weight. It can be specifically recognized by His McAb.