乙肝e抗原在大肠杆菌中的高效表达和优化  

Prokaryotic Expression of HBeAg and Its Optimal Condition

作　　者：周倩[1] 何小维[1] 彭运平 刘晓云 王海龙[1] 

机构地区：[1]华南理工大学轻工与食品学院,广东广州510640 [2]广州万孚生物技术有限公司,广东广州510640

出　　处：《现代生物医学进展》2009年第11期2098-2101,共4页Progress in Modern Biomedicine

基　　金：广东省部产学研结合项目(2006D90504011);广东省科技攻关项目(2006B40101013)

摘　　要：目的:探讨乙型肝炎病毒e抗原基因在大肠杆菌中进行高效融合表达,并对表达条件进行优化和影响因素的分析。方法:从乙肝患者阳性血清中提取DNA,设计引物扩增目的基因,按常规方法克隆入pET-GST载体谷胱甘肽-S-转移酶(GST)基因的下游,获得的重组质粒转化大肠杆菌BL21,用IPTG诱导目的基因片段的表达,经超声处理后SDS-PAGE电泳。对表达条件进行正交试验优化,并分析其影响因素的大小。结果:HBeAg以包涵体形式表达出来,其表达的GST融合蛋白的分子质量大约为44KD。最佳的表达条件为温度为30℃,IPTG浓度为0.6mmol/L,诱导时间为2h。其影响因素大小依次为诱导温度>诱导时间>IPTG浓度。结论:乙肝e抗原基因在大肠杆菌中获得了高效表达。Objective： To express HBeAg gene in E.coli efficiently, optimize its expression conditions and analyze the influential factors. Methods： The HBeAg gene was amplified by PCR from sera of the patient with HBV infection. Correspondent down-lead was designed to extend the aim gene. According to the previous method, they were sub-cloned into the downstream of GST gene in expression plasmid pET-GST. The recombinant vector was transformed into E.coli BL21, and GST fusion protein was expressed at the induction of IPTG. SDS-PAGE was used to identify the protein. The expression condition was oPtimized by orthogonal test and the influential factors analyzed. Results： HBeAg was expressed with form of inclusion bodies. The fusion proteins were 44KD.The optimal expression condition was 30℃, IPTG concentration 0.6mmol/L, induction time 2 hours. The ranking of the influential factors was： induction temperature 〉 induction time 〉 IPTG concentration. Conclusions： HBeAg genes were expressed in E. coli efficiently.

关 键 词：乙型肝炎病毒 乙型肝炎病毒E抗原 融合表达 正交试验 

分 类 号：R512.62[医药卫生—内科学]