猪环曲病毒RT-PCR检测方法的建立及其应用  

Development and application of RT-PCR assay for detection of porcine torovirus

作　　者：周璐[1] 李淞[1] 徐薇薇[1] 周远成[1] 赵睿[1] 蔡雨函[1] 吴江江[1] 朱玲[1,2] 徐志文[1,2] 郭万柱[1,2] 

机构地区：[1]四川农业大学动物生物技术中心,四川雅安625014 [2]动物疫病与人类健康四川省重点实验室,四川雅安625014

出　　处：《中国兽医科学》2012年第12期1264-1267,共4页Chinese Veterinary Science

基　　金：四川省科技支撑计划项目(2012NZ0001)

摘　　要：为建立猪环曲病毒快速检测方法,根据GenBank中猪环曲病毒S基因设计合成1对特异性扩增引物,建立了检测猪环曲病毒的RT-PCR方法。以伪狂犬病病毒、猪嵴病毒、猪传染性胃肠炎病毒、猪流行性腹泻病毒、A群猪轮状病毒、猪瘟病毒、猪链球菌和大肠杆菌进行特异性试验;结果显示,仅猪环曲病毒阳性模板扩增得到451bp的目的基因,测序结果与GenBank中猪环曲病毒S基因序列(GenBank登录号:AJ575372.1)的同源性为93%。敏感性试验显示,该方法能检测到的最低核酸含量为10pg。利用该方法对采集自四川省部分地区的43份临床样品进行检测;结果,临床样品中猪环曲病毒阳性率为25.6%。结果表明,建立的RT-PCR方法特异性强、灵敏度高,可用于猪环曲病毒的临床检测和流行病学调查。In order to establish a rapid RT-PCR assay for detection of porcine torovirus,a pair of special primers was designed according to the published sequences of porcine torovirus in GenBank.In the specificity test,we used this method to amplify pseudorabies virus,porcine kobuvirus,TGEV,PEDV,PRoV,CSFV,Streptococcus suis,and Escherichia coli.The results indicated that only porcine torovirus was successfully amplified a target fragment of 451 bp in size.When the sequence from the PCR products were compared to the sequence from GenBank（GenBank accession number：AJ575372）,they showed 93% identity.Testing of the sensitivity of RT-PCR indicated that the lowest detection limit was 10 pg nuclear acids.Forty three stool specimens collected from different farms in Sichuan Province were detected by the established method,and the positive rate was 25.6%.The results showed that the RT-PCR was specific and sensitive to detect porcine torovirus,which can apply for the clinic detection and molecular epidemiological survey.

关 键 词：猪环曲病毒 RT-PCR 检测 

分 类 号：S852.651[农业科学—基础兽医学]