表达传染性支气管炎病毒S1基因与鸡IL-18基因重组禽痘病毒的构建  

Construction of recombinant fowlpox virus coexpressing S1 of infectious bronchitis virus and chicken IL-18

作　　者：管倩[1,2] 陈红英[1] 崔保安[1] 杨明凡[1] 高文明[1,2] 李双亮[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]河南牧翔动物药业有限公司,河南郑州450008

出　　处：《中国兽医学报》2010年第11期1435-1439,共5页Chinese Journal of Veterinary Science

基　　金：国家"十一五"科技支撑计划资助项目(2006BAD06A08)

摘　　要：将含痘病毒启动子LP2EP2的鸡传染性支气管炎病毒S1糖蛋白基因和鸡IL-18(ChIL-18)基因同时插入到禽痘病毒转移载体pSY681中,获得重组禽痘病毒转移载体pSYS1/ChIL-18。用脂质体将其转染已感染禽痘病毒S-FPV-017株的鸡胚成纤维细胞,使其在鸡胚成纤维细胞内与禽痘病毒基因组发生同源重组,产生表达S1和鸡IL-18的重组禽痘病毒(rFPV-S1-ChIL-18)。在含有X-gal的营养琼脂培养基上进行蓝斑筛选,对重组病毒rFPV-S1-ChIL-18进行多次蚀斑克隆。以重组禽痘病毒DNA为模板,利用S1基因和鸡IL-18基因特异引物进行PCR,分别扩增出1条约1.7 kb和1条约0.6 kb的带。经间接免疫荧光法和T细胞转化试验证实感染rFPV-S1-ChIL-18的CEF中存在表达的S1和鸡IL-18。In order to develop recombinant fowlpox viruses（rFPV） coexpressing chicken IL-18（ChIL-18） and infectious bronchitis virus（IBV） S1 gene,S1 gene and chicken IL-18 were inserted into fowlpox virus transferring vector to construct pSYS1/ChIL-18.Recombinant expression plasmid pSYS1/ChIL-18 was transfected by lipofectamine on the chicken embryo fibroblasts cell（CEF） pre-infected with S-FPV-017.By selection of blue plaques on the CEF overlaid with agar containing X-gal,recombinant fowlpox virus（rFPV-S1-ChIL-18） were obtained and identified by PCR.The results indicated that recombinant fowlpox viruses contained chicken IL-18 and S1 genes and had the stable genetic properties.The expression of S1 was detected by indirect immunofluorescence in the recombinant virus-infected CEF.The expression of chicken IL-18 was detected by MTT in the recombinant virus-infected CEF.Development of recombinant fowlpox virus lay a foundation for study of biological function of expressed products and development of recombinant fowlpox viruses（rFPV） coexpressing chicken IL-18 and other protective antigen gene.

关 键 词：传染性支气管炎病毒 S1基因 鸡白细胞介素-18 重组禽痘病毒 

分 类 号：S852.65[农业科学—基础兽医学]