pSecTag2A-HBV-L真核分泌表达载体的构建及表达

Construction and Expression the Secreted Euraryotic Expression Vector of pSecTag2A-HBV-L

目的探索乙型肝炎病毒大蛋白（L蛋白）的分泌表达。方法用PCR方法克隆得到HBV-L基因，通过酶切、连接、转化构建pSecTag2A—HBV—L分泌型真核表达载体：通过脂质体介导的方法瞬时转染293T细胞，用RT-PCR和Westblot鉴定融合蛋白的表达。结果成功构建真核分泌表达载体pSecTag2A—HBV—L．HBV—L在mRNA和蛋白水平表达成功。结论融合IgKV_T2_C区域的外源性分泌信号能有效的引导L蛋白的组装和分泌．为进一步研究其DNA疫苗及免疫保护效果奠定了基础。

Objective To explore the secreted expression of hepatitis B virus large protein （L protein）. Methods The HBV-L gene was got by PCR, and it was constructed to the secreted eukaryotic expression vector of pSceTag2A-HBV-L by restrictive enzyme digestion, ligation and transformation. With pSecTag2A-HBV-L 293Tcells were transfected by liposome transfection, and the expression of fusion gene was analyzed by RT-PCR and West blot respectively. Results The eukaryotic expression vector named pSecTag2A-HBV-L was successfully constructed, and HBV-L protein could express successfully and demonstrate at HBV-L mRNA and protein levels. Conclusion HBV-L protein could be assembled and secreted effectively by the fusion of IgKV_T2_C exogenous secretion signal region, by which it could establish the foundation of the DNA vaccine and study the immune protective effect further.