结核分枝杆菌CFP10的基因克隆及表达

目的克隆结核分枝杆菌CFP10基因,在大肠杆菌中进行表达,为进一步研究其在结核病诊断中的价值奠定基础。方法以结核分枝杆菌H37Rv基因组DNA为模板,通过PCR方法对CFP10的基因进行扩增,以PET-30a(+)为载体构建重组质粒,将重组质粒先转化入大肠杆菌DH5α中,抽提质粒,进行双酶切鉴定,再转化到表达宿主菌BL21(DE3)中,以IPTG诱导表达,聚丙烯酰胺凝胶电泳(SDS-PAGE)分析蛋白表达形式。结果构建了具有正确基因序列的CFP10重组表达质粒,重组蛋白在大肠杆菌BL21(DE3)中以包涵体形式表达。结论目的基因克隆入宿主菌中并表达成功,为深入研究该蛋白的生物学、免疫学活性奠定了基础。

Cloning and expression of the gene coding for Mycobacterium tuberculosis antigen CFP10

To clone and express the gene coding for Mycobacterium tuberculosis antigen CFP10,and to establish a basis for diagnosis of tuberculosis,the gene coding CFP10 was amplified by polymerase chain reaction(PCR),and then the gene was inserted to vector pET-30a(+)to construct the recombinant plasmid.The recombinant plasmid was transformed into expressive vector E.coli BL21(DE3),and was induced with IPTG.The presence of recombinant protein in the expression vector was analyzed by SDS-PAGE.The recombinant CFP10 protein was expressed in inclusion body in E.coli BL21(DE3),and the target gene had been cloned into host bacterium.These results would establish a basis for diagnosis of tuberculosis.