金黄色葡萄球菌黏附素FnBPB-ClFA共表达质粒的构建与原核表达  

Construction and prokaryotic expression of the co-expression plasmid with FnBPB-ClFA gene encoding Staphylococcus aureus adhesin

作　　者：范鑫[1] 郝永清[1] 张爱荣[1] 史冬艳[1] 陈晓杰[1] 

机构地区：[1]内蒙古农业大学兽医学院,内蒙古呼和浩特010018

出　　处：《中国兽医科学》2011年第2期173-177,共5页Chinese Veterinary Science

摘　　要：采用PCR方法对编码金黄色葡萄球菌纤黏蛋白B的D区和纤黏蛋白原凝集素A的A区基因片段进行了特异性扩增,并通过重叠延伸PCR扩增了FnBPB-ClFA串联基因,构建了克隆质粒pMD19-FnB-PB-ClFA。将该基因片段定向插入到原核表达载体pET-32a(+)中,构建了表达质粒pET-FnBPB-ClFA,并将其转入宿主菌BL21(DE3)。SDS-PAGE分析表明,在1mmol/L IPTG诱导下,在51ku处出现了与目的蛋白一致的外源蛋白带。Western-blot分析表明,所表达的蛋白与目的蛋白一致。上述结果表明,该融合基因在原核细胞中成功表达。The genes encoding Fn-binding region D of fibronectin-binding proteins B and fibrinogen-binding region A of fibrinogen-binding proteins clumping factor A were amplified by PCR from Staphylococcus aureus chromosomal DNA and the amplified FnBPB-ClFA gene was cloned into pMD19-T vector by overlap extension PCR.The constructed plasmid pET-32a-FnBPB-ClFA was transformed into Escherichia coli BL21（DE3）.The bacterium was induced by 1 mmol/L IPTG and analyzed by SDS-PAGE and Western-blot.The approximately 51 ku exogenous protein was detected by SDS-PAGE.Western-blot indicated that the protein was the interested protein.The above results showed that the fusion protein was successfully expressed in prokaryotic cells.

关 键 词：金黄色葡萄球菌 纤维素结合蛋白B 凝集因子A 重叠延伸PCR 原核表达 

分 类 号：S852.611[农业科学—基础兽医学]