乙型肝炎病毒e基因在枯草杆菌细胞中的克隆和表达

乙型肝炎病毒e基因在枯草杆菌细胞中的克隆和表达 CLOING EXPRESSION OF HEPATITIS B VIRUS E GENE IN BACILLUS SUBTILIS CELLS 齐义鹏 Qi Yipeng Ma Xiaoweng Huang Yongxiu Tan Yeping 1 first-author 马晓雯 1 黄永秀 1 谭业平 1 武汉大学病毒学研究所 武汉大学病毒学研究所 Department of Virology,Wuhan University Department of Virology,Wuhan University 我们首先构建了一个枯草杆菌表达载体 pNQ122-tac,然后,在 tac 启动子下游插入乙型肝炎病毒（HBV）的含有 HBeAg 抗原决定簇和3′端的2/3编码序列的 Bgl Ⅱ片段.ELISA 检测证明,在枯草杆菌中,重组质粒 pHBe24能表达分子量为15.7 kD 的 HBV e 抗原,滴度高达1:12800,表达量为2.5 mg·L -1 ,占细菌总蛋白的8.86%,全部分泌到胞外,和 HBsAg、HmAg 都无交叉反应,兔抗-HBe 血清效价为1:1600. A HindⅢ-BamHI fragment with tac promoter from E.coli plasmid pDR540 has been inserted into the HindⅢ-BamHI site of Bacillus subtilis plasmid pNQ122.Further,a Bgl Ⅱ fragment carrying HBeAg gene was cloned into BamHI site dowmstream tac promoter.This recombinant DNA was transformed in B.subtilis As1176 by plasma-transform) resulting in selection of recombinant plasmid,such as pHBe24,containing HBeAg gene.The concentration of expression protein with molecular weight of 15.7 KDa produced by pHBe24 in B.subtilis cells is 2.5 mg·L -1 and 8.86% of total proteins in medium,in which expressive protein is secreted.It is shown with ELISA test that the expressive protein only contains HBeAg immunological activity （1:12800） and anti-HBe rabbit serum activity （1:1600）.We realized,therefore,HBV e gene is expressed in B.subtilis at first record. 枯草杆菌 tac 启动子 HBV e 基因 Bacillus subtilis tac promoter HBV e gene Q9;Q78;Q965.8 国家教委博士点基金;国家科委资助的课题;湖北省科委资助的课题 1993-04-01 2021-04-01 4