表达鸡传染性支气管病毒S1蛋白的重组假型杆状病毒的构建  

Construction of a Pseudotype Baculovirus Expressing S1 Protein of Infectious Bronchitis Virus

作　　者：罗琼[1] 樊惠英[1] 罗开健[1] 林文耀[1] 程晓亮[1] 任涛[1] 廖明[1] 

机构地区：[1]农业部动物疫病防控重点实验室,华南农业大学兽医学院,广东广州510642

出　　处：《华南农业大学学报》2010年第2期104-107,共4页Journal of South China Agricultural University

基　　金：“863”计划项目(2006AA10A205);教育部新世纪优秀人才支持计划(NCET-06-0752);国家自然科学青年基金(30800826);广东省博士启动基金(8451064201001131);农业微生物学重点实验室开发课题(20090010);华南农业大学校长基金(5500-K08240)

摘　　要：根据鸡传染性支气管炎病毒M41株的基因序列(GenBank:AY851295.1),设计特异性引物,利用RT-PCR方法扩增其S1基因,并将扩增产物克隆到杆状病毒转移质粒pFast-VSV-G-CMV中,获得重组质粒pFast-VSV-G-CMV-S1,进一步将该重组质粒转化到DH10Bac感受态细胞中,得到重组穿梭载体Bacm id-CMV-S1,再利用脂质体转染sf9昆虫细胞,获得重组杆状病毒Ac-V-S1.间接免疫荧光试验表明,该重组杆状病毒可以转导哺乳动物细胞并表达S1蛋白.According to the sequence of infectious bronchitis M41 （GenBank：AY851295.1） ,the S1 gene was amplified with specific primers and cloned the amplicon into the baculovirus transfer vector pFast- VSV-G-CMV to construct the recombinant plasmid pFast-VSV-G-CMV-S1. Then, the plasmid was transformed into Escherichia coli DH10Bac competent cells to obtain the recombinant shuttle vector Bacmid- CMV-SI after identifying the plasmid by digestion with restriction endonuclease and sequencing. Eventually, the recombinant Bacmid was transfected into sf9 ceils to produce the recombinant baculovirus Ac-V- S1 using the LipofectamineTM2000. Indirect immunofluoresent assay demonstrated the Ac-V-S1 could efficiently transduct the mammalian cells in vitro and the S1 gene was expressed successfully.

关 键 词：鸡传染性支气管炎 S1基因 重组假型杆状病毒 

分 类 号：S852.65[农业科学—基础兽医学]