赤羽病病毒套式RT-PCR检测方法的建立与初步应用  

Developmental and potential application of a nested RT-PCR for the detection of akabane virus

作　　者：张吉红[1] 黄素文[1] 闻伟刚[1] 李如松[1] 

机构地区：[1]宁波出入境检验检疫局,浙江宁波315012

出　　处：《畜牧与兽医》2010年第6期1-4,共4页Animal Husbandry & Veterinary Medicine

基　　金：宁波检验检疫局科研项目(NO:甬K06-2005)

摘　　要：根据GenBank中已发表的赤羽病病毒(AKAV)的S基因序列,设计了3条特异性引物,建立了检测AKAV的套式RT-PCR方法。特异性试验表明,该方法可以特异扩增出AKAV的S基因片段,但从牛病毒性腹泻病毒(BVDV)、传染性牛鼻气管炎病毒(IBRV)等13种对照病毒中均不能扩增出目的条带。敏感性试验表明,套式RT-PCR能够扩增10-5稀释度的病毒RNA(核酸含量约1.18 ng/μL),比普通RT-PCR高出1 000倍。用此方法检测130份奶牛血清样品与1份流产胎儿病料,均未检测到阳性样本,但AKAV在血清模拟样品中可被有效检出。研究结果表明,该方法灵敏、特异,为AKAV的检测提供了一个快速、有效的手段。A nested RT-PCR assay for detecting akabane virus(AKAV) was established using 3 primers.The primers were designed based on S gene information of AKAV in GenBank.The nested RT-PCR could only amplify the S gene fragment from AKAV,but not from 13 other viruses such as bovine viral diarrhoea virus(BVDV),infectious bovine rhinotracheitis virus(IBRV),and so on.The lowest detection limit for AKAV RNA was 10-5 dilution(RNA concentration was 1.18 ng/μL).The assay was 1 000 times more sensitive than normal RT-PCR.One hundreds and thirty cow serum samples and one sample from aborted fetus of dairy cow were tested by the nested RT-PCR for the detection of AKAV.The results showed that the above specimens were negative for AKAV.But in the mimic serum specimens,positive results could be obtained.It suggested that the nested RT-PCR was easy,specific and sensitive for detecting AKAV.It could be used for a rapid detection of AKAV.

关 键 词：赤羽病病毒 套式 RT-PCR检测 RT-PCR方法 初步应用 detection nested RT-PCR 牛病毒性腹泻病毒 牛鼻气管炎病毒 特异扩增 特异性引物 特异性试验 敏感性试验 血清样品 模拟样品 流产胎儿 基因序列 基因片段 核酸含量 GENBANK 

分 类 号：S8[农业科学—畜牧兽医]