虹鳟IPNV分离株VP2蛋白的表达及免疫原性检测  

Expression and immunogenicity analysis of VP2 protein of a Chinese infectious pancreatic necrosis virus from diseased rainbow trout Oncorhynchus mykiss

作　　者：赵景壮[1] 贺文斌[1] 徐黎明[1] 纪锋[1] 曹永生[1] 胡朝 卢彤岩[1] ZHAO Jing-zhuang;HE Wen-bin;XU Li-ming;JI Feng;CAO Yong-sheng;HU Chao;LU Tong-yan(Heilongjiang River Fisheries Research Institute,Chinese Academy of Fishery Sciences,Harbin 150070,China;Dujiangyan Administration of Sichuan Province,Dujiangyan 611830,China)

机构地区：[1]中国水产科学研究院黑龙江水产研究所,黑龙江哈尔滨150070 [2]四川省都江堰管理局,四川都江堰611830

出　　处：《大连海洋大学学报》2019年第2期179-185,共7页Journal of Dalian Ocean University

基　　金：中央级公益性科研院所基本科研业务费专项(2018HY-ZD0303);黑龙江省应用技术研究与开发计划项目(GA13B401);中国博士后科学基金资助项目(2018M630893)

摘　　要：为充分了解中国鱼类传染性胰腺坏死病病毒的系统进化,对流行病学调查分离出的虹鳟Oncorhynchus mykiss传染性胰腺坏死病病毒(IPNV)分离株进行了系统进化分析,利用软件综合分析了IPNV病毒表面蛋白VP2的跨膜区、亲水性、抗原性和表面可能性,对其特定区段进行了原核表达,并利用表达蛋白制备鼠抗血清及免疫学鉴定。结果表明:本实验室得到的分离株与中国已报道的毒株具有不同的来源;SDS-PAGE分析显示,表达纯化的VP2蛋白为单一条带,相对分子质量约为45 000;ELISA分析显示,利用表达蛋白制备的鼠抗血清与IPNV分离株细胞培养物的效价为1∶20 000,与VP2蛋白的效价为1∶40 000;间接免疫荧光分析显示,细胞感染IPNV病毒24、48 h后,该鼠抗血清均能与IPNV毒株发生特异性反应,并呈现出特异性绿色荧光。研究表明,本研究中表达的VP2蛋白与IPNV病毒的VP2蛋白具有相近的免疫原性,这一结果为IPNV检测方法的建立及疫苗的构建提供了重要的试验依据。An IPNV isolate obtained by epidemiological investigation from rainbow trout Oncorhynchus mykiss was analyzed in the evolution and transmembrane region,hydrophilicity,antigenicity and surface possibility of the VP2 protein in IPNV isolate by a software to understand the phylogenetic evolution of the Chinese infectious pancreatic necrosis virus and provide a theoretical basis for the virus detection and vaccine construction of the disease.Prokaryotic expression of the specific regions was carried out using pET-27b-VP2 vector and mouse antiserum was prepared with the expressed proteins.It was found that IPNV isolate was derived from different sources compared with the other isolates reported in China.SDS-PAGE revealed that the VP2 protein expressed and purified had a single band with relative molecular weight of approximate 45 000.ELISA analysis showed that the antisera against VP2 protein prepared here reacted specifically with both the recombinant VP2 protein and IPNV,with the maximal titer against recombinant VP2 protein of 1∶40 000,and the IPNV of 1∶20 000.Indirect immunofluorescence analysis(IFA)indicated that the antisera recognized IPNV in infected cells exposed for 24 h and 48 h via specific green fluorescence.The findings indicated that the antigenicity of recombinant VP2 protein was similar as that of IPNV,which provided important research basis for the detection and vaccine preparation of IHNV.

关 键 词：传染性胰腺坏死病病毒 蛋白表达 VP2蛋白 免疫原性 

分 类 号：Q785[生物学—分子生物学] S917[农业科学—水产科学]