原核载体表达的小反刍兽疫病毒N蛋白的保存期与相关抗原性、特异性研究  

Study on the Preservation Period,Antigenicity and Specificity of Peste Des Petits Ruminants Virus N Protein Expressed by Prokaryotic Vector

作　　者：孙雨[1] 宋晓晖[1] 肖颖 李秀梅 王睿男[1] 吕园园 蒋菲[1] 李晓霞 任雪建 王传彬[1] Sun Yu;Song Xiaohui;Xiao Ying;Li Xiumei;Wang Ruinan;LU Yuanyuan;Jiang Fei;Li Xiaoxia;Ren Xuejian;Wang Chuanbin(China Animal Disease Control Center,Beijing 102600,China;Chongqing Animal Disease Control Center,Chongqing 400000,China;Luoyang Modern Biotechnology Research Institute Co.,Ltd.,Luoyang,Henan 471000,China)

机构地区：[1]中国动物疫病预防控制中心,北京102600 [2]重庆动物疫病预防控制中心,重庆400000 [3]洛阳现代生物技术研究院有限公司,河南洛阳471000

出　　处：《中国草食动物科学》2020年第1期27-32,共6页China Herbivore Science

基　　金：国家重点计划研发项目（2016YFD0500901）;现代农业人才支撑计划（2130106-024)

摘　　要：为明确使用原核表达载体pET-32a表达纯化的小反刍兽疫病毒核衣壳蛋白(N)的最佳保存期,检测其相关抗原性与特异性,通过优化N抗原序列的密码子和蛋白表达纯化条件,在大肠杆菌表达系统中成功获得了3批次高效表达的小反刍兽疫病毒可溶性重组N蛋白。将3批次小反刍兽疫病毒N蛋白放置2~8℃保存,分别于1、2、3、6、9、12、15个月检验蛋白性状、蛋白浓度、纯度、抗原性和特异性等指标。经分析,各个时期蛋白的抗原浓度、抗原性、特异性等要素均符合标准,N蛋白保存15个月后,依然具有较强的检测抗原性与特异性。该研究结果为今后以小反刍兽疫N蛋白为基础建立相应的抗原捕获ELISA方法以及双抗原夹心ELISA抗体检测方法奠定了坚实的基础。In order to determine the optimum preservation time,the antigenicity and specificity of the nucleocapsid protein(N) of Peste des petits ruminants virus expressed and purified by prokaryotic expression vector p ET-32 a.Three batches protein of highly efficient E.coli expression system were obtained by optimizing the codon of N antigen sequence and optimizing the conditions of expression and purification. The soluble recombinant N protein of peste des petits ruminants virus was expressed. Three batches of N proteins were stored at 2-8 ℃ for 1,2,3,6,9,12 and 15 months respectively. The properties,protein concentration,purity,antigenicity and specificity of the proteins were tested. After analysis of protein concentration and other factors in each period,N protein still had strong antigenicity and specificity after 15 months of preservation. The results of this study lay a solid foundation for the establishment of corresponding antigen capture ELISA methods ruminants,competitive ELISA detection methods,indirect ELISA detection methods and double antigen sandwich ELISA antibody detection methods based on the N protein of pestilence petits in the future.

关 键 词：原核载体 小反刍兽疫病毒 N蛋白 保存期 抗原性 特异性 

分 类 号：S851.347.1[农业科学—预防兽医学]