鸟传染性支气管炎病毒类木瓜蛋白酶基因的克隆和表达  

Cloning and Expression of Papain-like Gene from Avian Infectious Bronchitis Virus

作　　者：袁宁[1] 王汉屏[1] 王艳[1] 

机构地区：[1]陕西教育学院生命科学系,陕西西安710061

出　　处：《安徽农业科学》2008年第18期7595-7596,7604,共3页Journal of Anhui Agricultural Sciences

基　　金：陕西省教育厅科研计划项目(07JK182)

摘　　要：[目的]为探究鸟传染性支气管炎病毒(IBV)的感染和复制过程及其防治提供信息和途径。[方法]以鸟IBV的cDNA为模板,利用PCR技术克隆IBV类木瓜蛋白酶基因。将回收片段与pGEX-6p-1载体连接并转入大肠杆菌DH5α中。在IPTG诱导下将克隆的重组质粒在大肠杆菌BL21中进行表达,经纯化获得目标蛋白。[结果]克隆的类木瓜蛋白酶基因全长为924 bp,编码308个氨基酸的完整开放读码框。它与Genbank中IBV M41毒株类木瓜蛋白酶基因的序列完全一致,证实该基因为IBV的类木瓜蛋白酶基因。在IPTG诱导下,BL21菌株成功表达目标基因和GST融合蛋白,融合蛋白约为60 kD。经纯化和酶切后所得蛋白产物的分子量约为34 kD。[结论]该基因的成功克隆和表达为有关蛋白结构与功能的研究奠定了基础。[ Objective] The research aimed to provide information and approach for probing into the infection and copy ,process of avian infectious bronchitis virus （IBV） and its control. [Method] With cDNA of avian IBV as template, papain-like genc of IBV was cloned by using PCR technology. The reclaimed fragment was connected with pGEX-6p-1 vector and introduced into Escherichia eoli DH5α. Through IPTG induction, the cloned recombinant plasmid was expressed in E. coli BL21 and the target protein was obtained after purification, [Result] Total length of the cloned papain-like genc was 924 bp and it coded a complete open reading frame with 308 amino acids. It was completely accordant with the sequence of papain-like genc in IBV M41 strain on GenBank website, which proved that this genc was papain-like gene of IBV. Through IPTG induction, the target gene and CST fusion protein were successfully expressed in BL21 strain and the fusion protein was about 60 kD. After purification and enzyme digestion, the obtained protein products was about 34 kD. [Conclusion] The successful cloning and exoression of this gene laid the foundation for studying the structure and functions of the related protein.

关 键 词：鸟传染性支气管炎 非结构蛋白 类木瓜蛋白酶 克隆 表达 SDS-PAGE 

分 类 号：S855.3[农业科学—临床兽医学]