猪传染性胃肠炎病毒重组N蛋白间接ELISA抗体检测方法的建立  

Development of an indirect ELISA for detecting antibody to transmissible gastroenteritis virus of swine using recombinant N protein

作　　者：徐向伟[1] 李彬[2] 孙冰[1] 郭容利[2] 俞正玉[2] 范宝超[2] 姜平[1] 倪艳秀[2] 何孔旺[2] 

机构地区：[1]南京农业大学动物医学院,江苏南京210095 [2]江苏省农业科学院兽医研究所,江苏南京210014

出　　处：《畜牧与兽医》2016年第5期31-38,共8页Animal Husbandry & Veterinary Medicine

基　　金：江苏省农业科技自主创新资金(CX(15)1056)

摘　　要：为建立一种检测猪传染性胃肠炎病毒(TGEV)血清抗体的间接ELISA方法,将TGEV的N基因片段克隆到p ET-28a载体中,转化至大肠杆菌BL21感受态细胞中进行诱导表达,并对表达的蛋白进行Western-blot鉴定。以纯化的重组N蛋白作为包被抗原,最终建立了检测TGEV抗体的间接ELISA方法。该ELISA方法与猪流行性腹泻病毒(PEDV)、猪瘟病毒(CSFV)、猪繁殖与呼吸系统综合征病毒(PRRSV)、口蹄疫病毒(FMDV)、猪圆环病毒2型(PCV2)等5种病毒阳性血清不发生交叉反应,表明建立的ELISA方法具有良好的特异性。本研究可为猪传染性胃肠炎的流行病学调查、诊断与防控奠定基础。T o establish an indirect ELISA method for the detection of serum antibody to porcine transmissible gastroenteritis virus of swine（ TGEV）,the N gene were amplified from TGEV genome and cloned into the p ET-28 a vector. The recombinant N proteincould be expressed in Escherichia coli BL21 by IPTG induction. The results of SDS-PAGE and Western blotting analysis revealed that the recombinant N protein was successfully expressed and had good reactivity with the antiserum to TGEV. Using the purified recombinant N protein as coating antigen,an indirect ELISA was developed for detection of antibody to TGEV. The recombinant N protein showed no cross-reaction with other five positive sera to PEDV,CSFV,PRRSV,FMDV and PCV2. It suggests that the indirect ELISA has good specificity. This study provides a basis for epidemiological investigation,diagnosis and control of TGEV infection.

关 键 词：猪传染性胃肠炎病毒 N基因 克隆 原核表达 间接ELISA 

分 类 号：S852.65[农业科学—基础兽医学]