牛病毒性腹泻病毒E2抗原表位表达及验证  

BVDV-E2 epitope analysis,expression and antigendetection

作　　者：黄美玲[1] 吴鹏[2] 李天森[2] 张国奇[1] 杨霞[2] 陈创夫[2] 盛金良[2] 

机构地区：[1]石河子大学生命科学学院,新疆石河子832000 [2]石河子大学动物科技学院,新疆石河子832000

出　　处：《生物技术》2016年第2期152-157,共6页Biotechnology

基　　金：国家国际科技合作专项项目("牛病毒性腹泻防治新技术的研究";No.2013DFR30970)

摘　　要：[目的]获得高效表达的重组牛病毒性腹泻病毒(Bovine viral diarrhoea virus,BVDV)E2免疫抗原蛋白。[方法]应用软件分析BVDV-E2蛋白抗原表位,将富含抗原表位基因片段连接表达载体p GEX-4T-1,并转化至大肠杆菌(DE3),经IPTG诱导表达,SDS-PAGE和Western Blot检测诱导表达蛋白,经镍柱分离纯化;以纯化蛋白为包被抗原,用间接ELISA检测抗原对免疫血清的反应性。[结果]成功克隆BVDV-E2抗原表位基因,并在大肠杆菌中高效表达,Western Blot和ELISA证实表达重组蛋白对BVDV阳性血清具有反应原性。[结论]获得大小为49.5 k Da的BVDV-E2抗原表位蛋白,并且该抗原稀释度在1:40,抗体稀释度在1:20时结合效果最佳。可用于后续的纳米抗体筛选的研究。[Objective]To get a high functionality antigenicity. [Methods]Predicting the epitope of BVDV- E2 protein by the gene sequences of BVDV- E2 in Gen Bank and online software. Then the expression vector（ p GEX- 4T- 1- E2） was constructed and cloned into the E. coli（ DE3）; Recombinant strains was induced to express by IPTG; expression products were analyzed by SDS- PAGE and Western Blot. Separation and purification of the proteins by nickel column. [Results]The gene sequences of the main antigen had been got,and the antigen proteins behaved a high binding capacity by Western Blot and it was also sensitive to the positive serum by ELISA analysis. [Conclusion]The expression of the BVDV- E2 protein epitopes was about 49. 5 k Da,it also had a best conbine effect at 1： 40 dilution of the antigen and 1∶ 20 dilution of the antibody,so it provides the foundation material for BVDV Nanobodies＇ Further research.

关 键 词：抗原表位 E2蛋白 Western BLOT 间接ELISA 

分 类 号：Q784[生物学—分子生物学]