一株携带EGFP基因的重组伪狂犬病毒变异株的构建  

CONSTRUCTION OF A RECOMBINANT PSEUDORABIES VIRUS VARIANT EXPRESSING EGFP

作　　者：童武[1] 郑浩[1] 梁超[1] 刘飞[1] 曹艳云[1] 李林[1] 田青[1] 郑旭晨[1] 单同领[1] 李国新[1] 童光志[1] 

机构地区：[1]中国农业科学院上海兽医研究所,上海200241

出　　处：《中国动物传染病学报》2015年第4期1-7,共7页Chinese Journal of Animal Infectious Diseases

基　　金：上海市自然科学基金项目(14ZR1448900);上海市科技兴农重点攻关项目(沪农科攻字(2015)第1-7号);中央科研院所公益性基础科研业务费项目(2014JB02);国家生猪现代产业技术体系项目(CARS-36)

摘　　要：本研究运用同源重组的方法,在伪狂犬病毒(Pseudorabies virus,PRV)变异株PRV JS-2012基础上,构建了一株携带增强型绿色荧光蛋白基因的标记重组伪狂犬病毒,命名为r PRV JS-2012-EGFP。经PCR方法鉴定及荧光显微镜观察分析,结果表明重组病毒构建成功,并能在感染细胞中稳定表达绿色荧光蛋白。生长曲线和空斑试验结果显示,r PRV JS-2012-EGFP与亲本株PRV JS-2012在体外具有相似的生长特性。将r PRV JS-2012-EGFP毒接种14日龄PRV阴性仔猪,能使实验猪100%发病,病死率高达40%,表明r PRV JS-2012-EGFP是一株具有强致病力的标记伪狂犬病毒变异株,在伪狂犬病毒变异株致病机制研究中具有一定的使用价值。An enhanced green fluorescent protein（EGFP） expression cassette was homologously recombined into the genome of a virulent Pseudorabies virus（PRV） variant strain PRV JS-2012, generating a recombinant PRV strain rPRV JS-2012-EGFP. The insert of EGFP gene were confirmed by PCR, and green fluorescence proteins were stably expressed in the cultured cells infected with rPRV JS-2012-EGFP by green fluorescence microscopy analysis. The assays of growth curve and plaque morphology revealed that rPRV JS-2012-EGFP showed similar virological properties to its parent strain PRV JS-2012. 14-day old piglets free of PRV antibodies were inoculated with rPRV JS-2012-EGFP. The morbidity of the infected piglets reached 100%, and the mortality reached 40%. These results showed that rPRV JS-2012-EGFP was a virulent markerPRV and could be utilized to study pathogenic characterization of PRV variant.

关 键 词：伪狂犬病毒 同源重组 重组病毒 绿色荧光蛋白 

分 类 号：S852.659.1[农业科学—基础兽医学]