口蹄疫病毒VPg2基因克隆表达及VPg2-ELISA检测抗体评价

 成功亚克隆了口蹄疫病毒（FMDV）太保毒株3ABC中VPg2基因，并将其插入pGEX-4T-1表达载体构建重组表达质粒，Western blotting分析表明，表达的VPg2蛋白与FMDV阳性血清发生反应。以VPg2表达蛋白为抗原建立间接ELISA方法，对背景清楚的试验牛血清进行检测，结果VPg2表达蛋白与空白对照组（C组）和灭活疫苗反复免疫组（CI组）牛血清均不发生反应，与未免疫直接攻毒组（D组）和免疫后攻毒组（I组）血清均发生反应；对D组和I组，VPg2表达蛋白抗体持续时间最长，可达90d，最早呈阳性反应时间为攻毒后第10天。VPg2表达蛋白抗体持续时间与先前测定的3ABC表达蛋白抗体几乎相同。用VPg2-ELISA方法检测2 170份进口阴性牛血清，12份出现假阳性，假阳性率为0.55%，而用3ABC-ELISA方法检测，48份出现假阳性，假阳性率为2.21%。

Cloning and Expression of VPg2 Gene Fragment of Foot-and-Mouth Disease Virus and the Evaluation of VPg2-ELISA for Antibody Detection

VPg2 gene fragment was successfully subcloned from 3ABC gene of the foot-and-mouth disease virus(FMDV) TaiBao strain and constructed VPg2 expressing plasmid by inserting the target gene fragment into pGEX-4T-1 vector.The expressed protein was analysed by Western blotting.The result showed strong reaction with FMDV positive serum.VPg2 protein ELISA was evaluated through testing sera of 4 cattle groups i.e.control group,vaccinated group,infected group and infected after vaccination group.Negative results were obtained for the control group and the vaccinated group.On the contrary,positive results were obtained for the infected group and part of the infected after vaccination group.In the infected group and infected after vaccination group,the duration of the antibody against VPg2 lasted for at least 90 days.The earliest detecting time of VPg2 antibody was 10th day after infection.The duration of the antibody against VPg2 was the same with the antibody against 3ABC tested previously.2170 negative sera were tested using indirect VPg2-ELISA and 3ABC-ELISA.The results showed that the false positive rates were 0.55% in VPg2 ELISA assay and were 2.21% in 3ABC-ELISA assay.