火鸡疱疹病毒荧光定量PCR检测方法的建立及初步应用  

DEVELOPMENT AND PRELIMINARY APPLICATION OF FLUORESCENCE QUANTITATIVE PCR FOR HERPESVIRUS OF TURKEY

作　　者：白银荣 张志飞 李雪松[2] 刘芹防[2] 陈鸿军[2] 杨健美[2] 李泽君[2] 滕巧泱[2] 张七斤[1] BAI Yin-rong;ZHANG Zhi-fei;LI Xue-song;LIU Qin-fang;CHEN Hong-jun;YANG Jian-mei;LI Ze-jun;TENG Qiao-yang;ZHANG Qi-jin(Inner Mongolia Agricultural University,Huhhot 010018,China;Shanghai Veterinary Research Institute,CAAS,Shanghai200241,China)

机构地区：[1]内蒙古农业大学,呼和浩特010018 [2]中国农业科学院上海兽医研究所,上海200241

出　　处：《中国动物传染病学报》2019年第1期8-12,共5页Chinese Journal of Animal Infectious Diseases

基　　金：国家自然科学基金项目(31472206);上海市科技兴农重点攻关项目(沪农科攻字(2015)第1-11号)

摘　　要：本研究针对火鸡疱疹病毒(Herpesvirus of turkey,HVT)FC-126毒株sorf1基因序列,设计特异性的扩增引物和探针,建立了检测HVT的TaqMan荧光定量PCR方法,并对其特异性、敏感性和重复性进行检测。结果表明,建立的荧光定量PCR方法灵敏度高,最低可检测到1×10~2 copies/μL,比普通PCR灵敏10倍;对鸭瘟病毒、传染性喉气管炎病毒、鹅细小病毒、4型禽腺病毒和8型禽腺病毒均无特异性扩增,具有较好的特异性;组间和组内变异系数均小于1%,表明该方法重复性好。用建立的荧光定量PCR方法对82份HVT样品进行检测,结果43份为阳性,普通PCR只检测出30份阳性样品。结果表明本研究建立的HVT TaqMan荧光定量PCR检测方法特异性好,灵敏度高,重要性好,为监测HVT的病毒含量提供了一种有效的手段。A pair of specific primers and a probe were designed for development of a fluorescence quantitative PCR according to the sorfi gene sequence of Herpesvirus of turkey (HVT) FC-126 strain. The results demonstrated that this PCR assay was more sensitive for HVT with a detection limit of 1 x l 〇2 copies/μL, which was 10-flod higher than conventional PCR. The assay also proved to be specific without amplification from Ehxck plague virus (DPV), Infectious laryngotracheitis vims (ILTV), Goose parvovirus (GPV), Fowl adenovirus serotype 4 (FAdV-4) and Fowl adenovirus serotype 8 (FAdV-8). The variabilities of inter- and intra-assay trials were less than 1%, indicating a good reproducibility. Subsequently, this fluorescent quantitative PCR was used to test clinical samples. As a result, 43 out of 82 samples were HVT positive in the fluorescent quantitative PCR while 30 samples were HVT positive in conventional PCR. Therefore, the fluorescent quantitative PCR developed in this study was highly specific and sensitive for detection of HVT.

关 键 词：火鸡疱疹病毒 荧光定量PCR sorf1基因 

分 类 号：S852.659.1[农业科学—基础兽医学]