鸡J亚群禽白血病病毒AH-J11株的全基因组感染性克隆构建  

Construction of Infectious Clone of Avian Leucosis Virus Subgroup J AH-J11 Strain

作　　者：王蓓[1] 刘芳[1] 王汉清[1] 李宁[1] 魏建忠[1] 刘光清[2] 王桂军[1] 

机构地区：[1]安徽农业大学动物科技学院,合肥230036 [2]中国农业科学院上海兽医研究所,上海200241

出　　处：《畜牧兽医学报》2013年第12期1976-1981,共6页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：安徽省科技攻关战略性新兴产业项目(11010302119);安徽省自然科学基金(11040606M90);公益性农业科研专项(201003012;0080319);"863"计划(2011AA10A200)

摘　　要：为探究鸡J亚群禽白血病病毒(ALV-J)的致病机理,采用PCR方法分4段分别扩增出J亚群禽白血病病毒安徽分离株AH-J11的前病毒cDNA,PCR产物经克隆鉴定,酶切后顺次连接,获得含有完整ALV-J AH-J11株前病毒cDNA的重组质粒,命名为pSK-AH-J11。将AH-J11株全基因序列克隆至pBluescriptⅡSK(+)载体中,得到重组质粒pBl-AH-J11,并将其转染鸡胚成纤维细胞(CEF)。通过间接免疫荧光试验,RT-PCR和Western blot检测,并经测序验证比对拯救前后病毒的gp85基因序列,结果均表明拯救出了1株重组J亚群禽白血病病毒(rALV-J)。本研究成功构建了鸡ALV-J的感染性克隆并救获了其重组病毒,为研究禽白血病的致病机理提供了良好的反向遗传操作技术平台。In order to explore the pathogenesis of subgroup J avian leukosis virus（ALV-J）, a full- length infectious clone of ALV-J （pSK-AH-Jll） was constructed by combining of four fragments using PCR method from AH-Jll isolated from broiler in Anhui province, then the complete ge- nome was cloned into pBluescript II SK（＋） vector. The recombinant plasmid pB1-AH-J11 was transfected into chicken embryo fibroblast （CEF） cells and the recombinant ALV-J was rescued. The rescued ALV-J （rALV-J） was identified by RT-PCR, Western blot, indirect immunofluores- cence assay and sequence comparision of gp85 gene with AH-J11strain, respectively. The results showed that the recombinant rALV-J was rescued in CEF. This study provides a useful platform for investigation of the pathogenesis and molecular biology of ALV-J.

关 键 词：J亚群禽白血病病毒 AH—J11株 全基因组 感染性克隆 病毒拯救 

分 类 号：S852.659.3[农业科学—基础兽医学]