马立克氏病病毒CVI988株病毒囊膜糖蛋白B启动子的克隆及活性分析  

Cloning Glycoprotein B(gB) Promoter of Marek's disease virus CVI988 Strain and Analysis of Its Transcriptional Function

作　　者：邱亚峰[1] 葛菲菲[1] 徐学清[1] 陈溥言[1] 

机构地区：[1]南京农业大学动物疫病诊断与免疫重点实验室,南京210095

出　　处：《农业生物技术学报》2006年第5期793-797,共5页Journal of Agricultural Biotechnology

基　　金：国家高技术研究与发展计划(863)项目(No.2002AA245051)资助

摘　　要：为了选择适宜的启动子调控外源基因的表达,以改善马立克氏病毒对载体的重组病毒的免疫保护力,克隆了马立克氏病病毒(MDV)CVI988株病毒囊膜糖蛋白B(gB)启动子,通过测序分析发现克隆的gB启动子与GenBank上发表的MDVGA株序列的同源性为99.5%。分别以β-半乳糖苷酶(β-galactosidase)和虫荧光素酶(luciferase)为报告基因,构建pSK-gB-LacZ和pgB-Luc真核表达载体。将pSK-gB-LacZ转染的中国地鼠卵巢(Chinesehamsterovary,CHO)细胞进行染色,出现蓝色,说明在CHO细胞中,MDVgB启动子可以有效地调控lacZ基因的转录。在CEF细胞中,利用虫荧光素酶系统将gB启动子与SV40及hCMV启动子进行活性比较,发现hCMV启动子的活性为gB启动子的31倍,SV40启动子为gB启动子的27倍。To improve the efficiency of the recombinant viruses in chickens with maternal antibodies, the glycoprotein B （gB） promoter of Marek＇s disease virus （MDV） CVI988 strain was cloned and compared with sequence of MDV GA strain in GenBank. The result showed that the homologue was about 99.5%. And a plasmid named pSK-gB-LacZ, containing Escherichia coli lacZ gene under control of MDV gB promoter, was constructed to conform the transcriptional function of gB promoter. At the same time, the other plasmid named pgB-Luc, containing luciferase gene under the control ofMDV gB promoter, was constructed to analyse the transcriptional activity of MDV gB promoter, pSK-gB-LacZ was transfected into Chinese hamster ovary （CHO） cell line by calcium phosphate, and the cells were stained by X-gal. The result showed that MDV gB promoter could successfully control the expression of the lacZ gene in CHO cell line. Moreover, pgB-Luc, pSV-Luc or phCMV-Luc were respectively cotransfected into chicken embryo fibroblast （CEF） with pSV-β-Gal to compare their transcriptional activity. The result indicated that the transcriptional activity ofhCMV promoter was 310% of that ofgB promoter, and the transcriptional activity of the SV40 promoter was 270% of that ofgB promoter.

关 键 词：马立克氏病毒 启动子 转录 转染 

分 类 号：S188[农业科学—农业基础科学]