牛病毒性腹泻病毒E^(rns)-E2融合蛋白在果蝇S2细胞中的表达与鉴定  

Expression and Identification of the Fusion Protein E^(rns)-E2 of Bovine Viral Diarrhea Virus in Drosophila S2 Cell

作　　者：孙宇[1,2] 史利军[1] 孙卫华[1] 章金刚[1] 

机构地区：[1]军事医学科学院野战输血研究所,北京100850 [2]解放军第305医院,北京100017

出　　处：《生物技术通讯》2008年第6期862-865,共4页Letters in Biotechnology

摘　　要：目的:利用果蝇S2细胞表达牛病毒性腹泻病毒(BVDV)Erns-E2融合蛋白,并对其抗体结合能力进行鉴定。方法:用RT-PCR方法扩增BVDV NADL株Erns和E2蛋白的编码基因,利用(G4-S)3柔性15肽基因将扩增的2个基因连接,再与昆虫表达载体pMT/BiP/V5-His连接构建重组表达载体pMT/BiP/V5-His-Erns-E2,将后者与筛选质粒pCoBlast共转染果蝇S2细胞后表达Erns-E2融合蛋白,并对表达产物进行鉴定。结果:SDS-PAGE结果表明,融合蛋白相对分子质量为76800;Western blotting检测表明,该融合蛋白具有与BVDV抗体良好的结合能力。结论:BVDV的Erns-E2融合蛋白能在果蝇S2细胞中进行表达;经鉴定,表达产物具有良好的抗体结合能力,可用于抗原检测。Objective： To expressE^rns-E2 fusion protein of bovine viral diarrhea virus（BVDV） by Drosophila S2 protein expressed system and to detect the antigenieity of the fusion protein. Methods： The genes endoding E^rns and E2 of BVDV NADL strain were amplified by RT-PCR, and were ligated by （G4-S）3 gene as fusion gene, then was cloned into pMT/BiP/V5-His vector for constructing the recombinant expression vector pMT/BiP/V5-His-E^rns-E2 Expression vector pMT/BiP/V5-His-E^-E2 and screen vector pCoBlast were cotransfected into s2 cells to generate fusion protein. The anti- genieity of expressed protein was identified by Western blotting. Results： A specific and simple band at about 76.8 kDa was showed by SDS-PAGE and the protein was proved to have good antibody binding capacity by Western blotting analysis. Conclusion： This study indicated that E^rns-E2 fusion gene of BVDV NADL strain can be effective translated in S2 cells as respected, and theE^rns-E2 fusion protein can be used as an envelope antigen for BVDV antibody detection.

关 键 词：牛病毒性腹泻病毒 E^rns蛋白 E2蛋白 融合表达 

分 类 号：Q78[生物学—分子生物学]