猪传染性胃肠炎病毒HN-2012分离株S基因的遗传变异分析及其真核表达  

Genetic variation analysis and eukaryotic expression of S gene of porcine transmissible gastroenteritis virus HN-2012 strain

作　　者：陈雅君[1,2] 程慧芳[1] 徐卫松[1] 刘中原[2] 寇亚楠[1] 张莉娟[1] 崔保安[1,2] 胡慧[1,2] 

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002 [2]河南省动物性食品安全重点实验室,河南郑州450002

出　　处：《中国兽医科学》2015年第1期32-36,共5页Chinese Veterinary Science

基　　金：国家自然科学基金项目(31101796);教育部博士点新教师基金项目(20104105120005);河南省基础与前沿技术研究计划项目(112300410161)

摘　　要：以猪传染性胃肠炎病毒(TGEV)HN-2012株的cDNA为模板,经克隆测序后,将得到的S基因序列及其推导的氨基酸序列与NCBI中不同来源的TGEV毒株的相应序列进行同源性和亲缘关系的比较分析,构建系统进化树进行遗传变异分析;然后将S基因亚克隆至真核表达载体pCAGGS-S-flag中,转染293T细胞进行表达,利用flag标签抗体进行Western-blot分析。结果显示,TGEV HN-2012株的S基因与其他毒株间核苷酸和氨基酸的同源性分别为95.1%~99.1%和93.6%~95.2%,与H株的亲缘关系较近,与我国其他毒株的亲缘关系较远。Western-blot分析中可见大小约为160ku的目的条带,表明S基因成功在293T细胞中表达。本试验结果为深入研究TGEV S蛋白的生物学功能奠定了基础。The S gene amplified from the porcine transmissible gastroenteritis virus(TGEV)HN-2012 strain by PCR using cDNA as template was cloned into the cloning vector and sequenced.Sequence analysis showed that the homology of nucleotide and amino acid sequence among different TGEV HN-2012 strains were from 95.1% to 99.1%,and from 93.6% to 95.2%,respectively.The isolated strain HN-2012 had closer correlation with the isolated strain H.The S gene was cloned into the eukaryotic expression vector pCAGGS-S-flag and transfected into 293 Tcells.The expression of S protein was verified by Western-blot.Using the flag label antibodies,a band of about 160 ku size was visible in the Western-blot test.The eukaryotic expression plasmid pCAGGS-S-flag with S gene from the strain HN-2012 was successfully constructed and expressed in 293 Tcells.The above-mentioned result laid the foundation for the study on biological functions of S protein of TGEV in the future.

关 键 词：猪传染性胃肠炎病毒 S蛋白基因 293T细胞 真核表达 

分 类 号：S852.651[农业科学—基础兽医学]