多联重组痘苗病毒的构建及鉴定  

Construction and selection of the recombinant vaccinia Tiantan strain with four exogenous genes combined

作　　者：李一权 兰添[2] 胡宁宁[2] 姜爽[2] 李敏[2] 曹亮[1,2] 曹佳媛 范园园[2] 李霄[2] 金宁一[2] 

机构地区：[1]吉林农业大学动物科学技术学院,吉林长春130118 [2]军事医学科学院军事兽医研究所,吉林长春130122

出　　处：《中国兽医学报》2016年第7期1081-1085,1091,共6页Chinese Journal of Veterinary Science

基　　金：吉林省重点科技攻关资助项目(20130206041NY);吉林省产业技术创新战略联盟项目(20140309006YY)

摘　　要：构建含有埃博拉出血热病毒(Ebola hemorrhagic fever virus,EHFV)小片段分泌蛋白S基因、马尔堡出血热病毒(Marburg hemorrhagic fever virus,MHFV)GP基因、辛诺柏病毒(Sin Nombre virus,SNV)G2糖蛋白基因及非洲欧尼恩病毒(Africa O’nyong nyong virus,O’NNV)E2蛋白基因的重组痘苗病毒。设计含EGFP筛选标记和4种外源基因的穿梭质粒,并利用同源重组技术、荧光筛选技术和Cre/LoxP敲除系统,最终获得四联重组痘苗病毒。使用PCR方法鉴定重组痘苗病毒及其遗传稳定性;用透射电镜观察病毒形态。结果显示:所构建的多联重组痘苗病毒rVTT-J^-4^+具有良好的遗传稳定性,并且具备痘苗病毒的完整形态。结果表明:成功构建了含有4种外源基因的重组痘苗病毒rVTT-J^-4^+,为后续多联疫苗的研究奠定了基础。To construct a recombinant vaccinia virus contained the Ebola hemorrhagic fever virus（EHFV）small fragments secreted protein S gene,Marburg hemorrhagic fever virus（MHFV）GP gene,Sin Nombre virus（SNV）G2 glycoprotein gene and Africa O＇nyong nyong virus（O＇NNV）E2 protein gene.Designed a shuttle plasmid containing the EGFP selectable marker and four exogenous genes.Constructed the recombinant vaccinia virus by using homologous recombination technique,fluorescence screening technique and Cre/LoxP system.Using PCR to identify recombinant vaccinia virus and its genetic stability.Virus morphology was identified by TEM.The recombinant vaccinia virus rVTT-J^-4~＋ has a good genetic stability and a complete morphology.We successfully constructed the recombinant virus rVTT-J^-4~＋ with four exogenous gene,and laid the foundation for the subsequent multi-vaccine research.

关 键 词：多联疫苗 痘苗病毒 同源重组 EGFP 

分 类 号：S852.65[农业科学—基础兽医学]