伪狂犬病病毒gE基因主要抗原表位区的原核表达及间接ELISA方法的建立  

Prokaryotic expression of main antigen epitope of pseudorabies virus gE gene in Escherichia coli and establishment of indirect ELISA

作　　者：吉艺宽[1] 王雨[1] 程艺[1] 张宝石[1] 向柯宇 琚春梅[1] 

机构地区：[1]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》2015年第4期11-15,共5页Journal of South China Agricultural University

基　　金：国家自然科学基金(31001074)

摘　　要：【目的】为了区分伪狂犬病病毒疫苗免疫猪和自然感染猪,建立快速诊断的方法.【方法】利用PCR方法从伪狂犬病病毒中扩增出糖蛋白E(g E)基因的主要抗原表位区,用Bam HⅠ和HindⅢ双酶切后,插入原核表达载体p ET-32a(+)中,构建重组表达质粒p ET-g E184,并转化至大肠埃希菌Escherichia coli BL21(DE3),经IPTG诱导后进行SDS-PAGE电泳和Western-blot检测,用纯化后的g E184蛋白作为包被抗原,建立间接g E184-ELISA检测方法.【结果和结论】用该方法检测290份临床猪血清样本,并与商品化ELISA试剂盒检测结果比较,两者的符合率为93.1%.【Objective】To distinguish pseudorabies virus（ PRV）-infected pigs from those vaccinated with glycoprotein E（ g E）-negative vaccine,the method of rapid diagnosis was established. 【Method】The g E gene fragment of PRV containing the main antigen region was amplified by PCR and cloned into the prokaryotic expression vector p ET32a（ ＋） to establish the recombinant plasmid p ET-g E184. The recombinant plasmid was transformed into Escherichia coli BL21（ DE3）. After being induced with IPTG,the target protein was identified by SDS-PAGE and Western-blot. Wells of ELISA plates were coated with purified recombinant protein g E184 to establish the indirect g E184-ELISA. 【Result and conclusion 】Two hundred ninety clinical porcine sera samples have been detected by this ELISA. Compared with commercial ELISA Kit,the coincidence rate was 93. 1%.

关 键 词：伪狂犬病毒 GE基因 抗原表位区 原核表达 gE184-ELISA 

分 类 号：S852.65[农业科学—基础兽医学]