狂犬病病毒快速荧光RT-PCR检测方法研究与应用  

Studies and application of the fast detection rabies virus (RV) by real time RT-PCR

作　　者：高志强[1] 张鹤晓[1] 柏亚铎[1] 赖平安[1] 谷强[1] 凌凤俊[1] 张伟[1] 张向东[1] 段生涛[1] 

机构地区：[1]北京出入境检验检疫局,北京通州101113

出　　处：《中国兽医杂志》2010年第1期14-17,共4页Chinese Journal of Veterinary Medicine

基　　金：北京市科技计划:国家科技计划项目衔接-科技奥运专项(Z07001000560715);国家质量监督检验检疫总局科技计划(2007IK027)

摘　　要：采用TaqMan方法,根据古典狂犬病病毒非编码区及N基因编码区序列,设计合成多对引物和多条探针,通过对引物、探针的筛选,反应条件的选择和优化,建立了古典狂犬病病毒荧光RT-PCR检测技术。与病毒分离鉴定、常规RT-PCR试验比较表明,建立的荧光PCR检测技术快速、敏感,检测时限3 h以内。特异性试验表明本方法与犬瘟热病毒,犬细小病毒,犬Ⅰ型腺病毒不发生交叉反应。初步动物感染试验及定量检测研究及表明,本方法可用于感染动物不同组织样品中病毒的相对定量。对132份临床样品和4种商品化疫苗进行检测研究表明,本方法适用于样品中狂犬病病毒的直接检测以及疫苗中活病毒或灭活病毒的检测。In this study, a real-time RT-PCR（RRT-PCR） was developed to detect classical rabies virus （RV） using Taq Man technique by the selection and optimization of primers, probes and reaction conditions. Compared with virus isolation（VI） and normal RT-PCR, the developed RRT-PCR was fast and sensitive, with the detection time less than 3 hours. The results also clearly demonstrated high specificity of developed RRT-PCR for the detection of RV by testing canine distemper virus, canine parvovirus and ca- nine adenovirus type 1. Furthermore, primary animal infection and quantity tests showed RRT-PCR fit for virus relative quantification from samples from different tissues. The results from the examination of 132 clinical samples and 4 kinds of merchandised vaccine suggested that RRT-PCR could directly detect RV from samples and from killed or modified vaccines.

关 键 词：狂犬病病毒 荧光RT—PCR 研究与应用 

分 类 号：S852.659.5[农业科学—基础兽医学]