HBV中片段表面抗原及E抗原真核表达质粒的构建与体外表达

Construction and in Vitro Expression of Eukaryotic Expression Plasmids of HBV Middle Envelope Protein and HBV E Antigen

目的 构建乙型肝炎病毒 ( HBV)中片段表面抗原真核表达质粒和 E抗原真核表达质粒。方法 将编码 HBV中片段表面抗原 ( pre S2 +S)的基因片段和 E抗原 ( HBe Ag)的基因片段分别插入真核细胞表达载体 ( p JW430 3)中。构建成的重组体经筛选鉴定后 ,再转染哺乳动物细胞 ,检测表达产物。结果 电泳鉴定显示目的基因片段正确插入了表达载体。经筛选、纯化的阳性克隆转染哺乳动物细胞后可表达 pre S2 +S和 HBe Ag。结论 HBV中片段表面抗原真核表达质粒 ( p JW430 3/HBV- S2 .S)和 E抗原真核表达质粒 ( p JW430 3/ HBe)

Objective To construct the eukaryotic expression plasmids of HBV middle envelope protein and HBV E antigen. Methods Two recombinant plasmids were constructed by inserting HBV middle envelope protein gene fragment or E antigen gene fragment into the reading frame of plasmid pJW4303. The positive clones were screened and identified by restrictive endonuclease digestion. Transient expressions were performed by transfection of 293T cells with purified recombinants. The expressive products were detected by ELISA. Results The electrophoresis of products with the endonuclease digestion showed that the target gene fragments have been inserted into pJW4303. The expression of HBV middle envelope protein and HBeAg were demonstrated by ELISA in cell culture supernatant and/or cell lysis of 293T cells. Conclusion The results showed that the eukaryotic expression plasmids of pJW4303/HBVS2.S and pJW4303/HBe were constructed successfully.