猪嵴病毒RT-PCR检测方法的建立及应用  

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

作　　者：李淞[1,2] 朱玲[1,2] 周远成[1,2] 吴云飞[1,2] 陈蕾[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620514 [2]四川农业大学动物疫病与人类健康四川省重点实验室,四川雅安620514

出　　处：《中国兽医学报》2013年第8期1150-1153,共4页Chinese Journal of Veterinary Science

基　　金：四川省科技支撑计划资助项目(2011FZ0065);四川省科技支撑计划资助项目(2012NZ0001)

摘　　要：根据GenBank中猪嵴病毒(porcine kobuvirus)3D-RNA聚合酶基因序列设计并合成了1对特异性引物,建立了一种猪嵴病毒RT-PCR检测方法。反应产物测序结果与GenBank中猪嵴病毒SH-W-CHN/2010/China株比较,基因序列同源性为93%。利用该方法对猪蓝耳病毒、猪伪狂犬病病毒、猪瘟病毒、猪传染性胃肠炎病毒、猪流行性腹泻病毒、猪轮状病毒、链球菌、猪巴氏杆菌和大肠杆菌等病原核酸进行扩增,均无条带出现,表明该方法具有较高的特异性;敏感性试验表明,该方法最低能检测到的病毒核酸含量为1pg。利用所建立的RT-PCR方法检测采集自我国四川地区123份临床样品,结果阳性率为59.3%,表明猪嵴病毒在四川地区猪群中流行率较高。In order to establish a RT-PCR assay for detection of porcine kobuvirus,a pair of primers were designed from conserved regions according to the published 3D-RNA ploymerase gene se- quences in GenBank. The RT-PCR diagnostic method for porcine kobuvirus was established base on the optimization of RT-PCR reaction conditions. Sequence analysis of PCR product showed that the homology of gene sequence is 93 percent like other porcine kobuvirus（SH-W-CHN/2010/ China）. We used this method to amplify PRRSV, PRV, CSFV, TGEV, PEDV, PoRV, Streptococcus suis,Pasteurella multocida ,Escherichia coli ,the results proved that this RT-PCR assay has high specificity. The sensibility of RT-PCR indicated the lowest of nucleic content is 1 pg for detection. Using the established assay detected 123 stool specimens collected from different farms in Sichuan province,the positive rate is 59.3% ,it seems that porcine kobuvirus is very prevalent in China.

关 键 词：猪嵴病毒(porcine kobuvirus) 3D-RNA聚合酶 RT-PCR 

分 类 号：S852.65[农业科学—基础兽医学]