马立克病病毒gB主要抗原决定簇融合基因的真核表达  

Construction and Expression of Eukaryotic Plasmid of the Fused Gene Containing the Major Antigen Cluster of Marek's disease virus Glycoprotein B and HA Epitope

作　　者：葛菲菲[1] 刘佩红[1] 王建[1] 周锦萍[1] 沈莉萍[1] 徐锋[1] 孙泉云[1] 

机构地区：[1]上海市畜牧兽医站,上海201103

出　　处：《动物医学进展》2007年第9期26-30,共5页Progress In Veterinary Medicine

摘　　要：为了合成由HA表位和串联的马立克病病毒（MDV）gB主要抗原决定簇基因所构成的融合蛋白基因,并在真核中表达,选取了MDV GA株gB 250位～460位氨基酸为目的片段,应用基因重组技术,通过一个由柔性氨基酸组成的linker,使该基因片段在体外串联,并在串联片段的前端加上HA表位,末端加上终止密码子,以形成完整的阅读框;通过测序分析证明,引入的HA表位序列以及融合基因的阅读框完全正确.将融合基因克隆入真核载体pIRES中,从而获得真核表达质粒pCMV-HAgB（L）,利用脂质体将该表达质粒转染CHO细胞,通过间接免疫荧光（IFA）和RT-PCR鉴定,发现该融合基因可以在真核细胞中正确表达,为开发新型的马立克病疫苗奠定了基础.In order to construct eukaryotic expression vector of a fused gene containing HA epitope and the major antigen cluster of Marek＇s disease virus glycoprotein B in eukaryotic cell, the 250 - 460 amino acids of the glycoprotein B of Marek＇s disease virus GA strain was selected as target fragment. The target fragment were tandem ligased in vitro through a flexible linker by recombinant DNA techniques. On the other hand, the HA epitope gene and TAA was fused to 5＇ and 3＇ of the tandem fragment separately to obtain the right open reading fragment （ORF） for gene expression. Sequence analysis indicated that the HA epitope gene and ORF were accurate. Then, the fused gene was cloned into pIRES to obtain pCMV-HAgB （L）. pCMV-HAgB（L） was transfected into CHO by use of Lipofectin 2000. The result of IFA and RT- PCR indicated that the fused gene was expressed. The research lay a favorable foundation for developing the novel MDV vaccine.

关 键 词：真核表达 马立克病病毒 抗原决定簇 转染 

分 类 号：Q785[生物学—分子生物学]