截短的乙型肝炎表面抗原的原核表达及其特性的研究

目的 在原核系统中表达截短的乙型肝炎表面抗原(HBsAg),并检测其抗原特性。为乙型肝炎疫苗(主要是治疗性疫苗)免疫原的选择进行初步的研究和探讨。 方法 利用聚合酶链反应(PCR)特异性扩增编码HBsAg羧基末端152和124个氨基酸的基因片段,均定向克隆至质粒pET32a( ),并将重组质粒分别转染至大肠杆菌(BL21)。 结果 重组质粒均成功构建。大肠杆菌表达产物氨基末端有1个含6个组氨酸肽段的150个氨基酸融合部分,以包含体形式存在。用SDS-PAGE、Western blot、ELISA等方法对原核表达产物进行检测,结果表明原核表达产物具有HBsAg反应原性。 结论 截短的HBsAg能在原核系统中表达,可为后续研究做准备。

Prokaryocytic expression of the shortened hepatitis B surface antigen and antigenic analysis

Objective To study the expression of the shortened hepatitis B surface antigen in prokaryocyte and detect the antigenic characters. Methods Firstly, the gene fragments coding the 152 and 124 amino acids of the carboxyl terminus of hepatitis B surface antigen (HBsAg) were amplified by polymerase chain reaction (PCR). Secondly, they were cloned to plasmid pBKS , and the accuracy of those constructions were confirmed by restriction enzyme digestion and DNA sequencing. Then, they were cloned to prokaryocytic expression vector-plasmid pET32a( ). The recombinant plasmids were transfected into E.coli BL21 and induced to express with IPTG. Results The recombinant plasmids were successfully constructed. In E.coli BL21, the protein was expressed in a fusion fashion and could be recognized by monoclonal antibody against HBsAg with ELISA and Western blot. Conclusion The shortened HBsAg can be expressed in prokaryocyte.