鸡传染性喉气管炎病毒TaqMan real-time PCR检测方法的建立  

Development of a TaqMan real-time PCR for detection of infectious laryngotracheitis virus

作　　者：赵妍[1] 孔聪聪[1] 张晓敏[2] 崔红玉[1] 石星明[1] 赵晓岩[1] 薛美[1] 胡顺磊[1] 闫帅[1] 牛秀杰[1] 李巧玲[1] 王玫[1] 王云峰[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/禽传染病研究室,黑龙江哈尔滨150001 [2]云南农业大学预防兽医系,云南昆明650201

出　　处：《中国预防兽医学报》2012年第8期642-646,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：中央级公益性科研院所基本科研业务费项目(0302012009)

摘　　要：为建立鸡传染性喉气管炎病毒(ILTV)TaqMan Real-time PCR检测方法,本研究根据GenBank中登录的ILTV gB基因序列设计了2对引物与一条特异性TaqMan探针,通过对反应体系和反应条件的优化,特异性、敏感性以及重复性试验,证明该方法在核酸含量108拷贝/μL～101拷贝/μL范围内具有良好的线性关系;能够检测初始模板中10-3EID50的病毒核酸及16拷贝的标准品;与其它相关的鸡源病毒均无交叉反应,并且批内、批间变异系数均小于2%,具有良好的重复性。该检测方法的建立为ILTV的临床检测和定量分析提供了一种快速、准确的技术手段。Infectious laryngotracheitis virus （ILTV） is the causative agent of a highly contagious upper respiratory tract infection in chickens. To develop a sensitive method for ILTV detection, a TaqMan real-time PCR assay was established using two pairs of primers and a specific probe designed according to the conserved region of ILTV gB gene. Under the optimized reaction conditions, the test results showed that this method was specific for detecting ILTV with a detection limit of 10^-3 EID50, and no cross-reactions to other chicken virus. The repeatability tests indicated that the inter- and intra-variation were less than 2%. The development of this rapid and sensitive detection method could be applied in clinical diagnosis of ILTV infections and quantitative analysis of ILTV.

关 键 词：鸡传染性喉气管炎病毒 GB基因 TAQMAN REAL-TIME PCR 

分 类 号：S852.65[农业科学—基础兽医学]