禽腺病毒血清4型纳米PCR检测方法的建立与初步应用  

Establishment and application of Nano PCR assay for detection on Fowl aviadenovirus serotype 4

作　　者：王利丽[1,2,3] 郑丽 李富强[1,2,3] 李秀丽 任卫科[1,2,3] 路超 张莉[1,2,3] 董志民 鄢明华[1,2,3] WANG Li-li;ZHENG Li;LI Fu-qiang;LI Xiu-li;REN Wei-ke;LU Chao;ZHANG Li;DONG Zhi-ming;YAN Ming-hua(Tianjin Animal Husbandry and Veterinary Research Institute,Tianjin 300381,China;Tianjin scientific observation experiment station for veterinary medicine and diagnosis technology,the ministry of agriculture of China,Tianjin 300381,China;Tianjin Engineering Research Center for Livestock and Poultry Health Breeding,Tianjin 300381,China.)

机构地区：[1]天津市畜牧兽医研究所,天津300381 [2]农业部兽用药物与诊断技术天津科学观测实验站,天津300381 [3]天津市畜禽健康养殖技术工程中心,天津300381

出　　处：《中国预防兽医学报》2019年第7期701-704,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家重点研发计划项目(2016YFD0500800);天津市企业科技特派员项目(18JCTPJC67800)

摘　　要：为建立一种快速有效检测禽腺病毒血清4型(FAdV-4)的方法,本研究根据FAdV-4五邻体(Penton)基因序列,设计合成1对特异性引物,经过条件优化,建立了检测FAdV-4的纳米PCR方法(Nano PCR)。特异性试验结果显示,该方法对鸡新城疫病毒等12种家禽常见病原及国内报导的FAdV-1、FAdV-2、FAdV-8a、FAdV-8b、FAdV-11等5个主要血清型病毒的扩增结果均为阴性,仅FAdV-4检测结果为阳性,特异性较强。敏感性试验结果显示,该方法对FAdV-4的最低核酸检出量为54.0拷贝/μL,较常规PCR方法高10倍,敏感性较高。应用该方法和病毒分离培养法同时对87份临床样品进行检测,二者均检出14份阳性样品,总体符合率为100%。以上结果表明本研究建立的Nano PCR方法特异性好、敏感性高,可以用于FAdV-4的临床检测和分子流行病学调查。To establish a rapid,specific and sensitive Nano PCR detecting method for Fowl aviadenovirus serotype 4(FAdV-4)diagnosis,a pair of primers were designed based on Penton gene of FAdV-4.The Nano PCR method was developed after the optimization of amplification conditions.The specific detection results of 12 common poultry pathogens and the FAdV-1,FAdV-2,FAdV-8a,FAdV-8b,FAdV-11 were all negative except for FAdV-4.The detection limit of the viral nucleic acid was 54.0 copies/μL.The Nano PCR method was approximately 10-fold more sensitive than regular PCR assay.Among 87 clinical samples,14 positive samples were identified by virus isolation and the Nano PCR method.The overall coincidence rate of two methods was 100%.These results indicated that the Nano PCR has good specificity,sensitivity and repeatability,which can be applied widely in clinical diagnosis and field surveillance of FAdV-4 infection.

关 键 词：禽腺病毒血清4型 五邻体基因 NanoPCR检测 

分 类 号：S852.65[农业科学—基础兽医学]