鸭甲型肝炎病毒3型和鸭圆环病毒双重荧光定量PCR检测方法的建立  

Development of a Duplex Real-time PCR for Detection of Duck Hepatitis Virus Type 3 and Duck Circovirus

作　　者：于新友 李天芝 苗立中[2] 谢金文[2] 王文秀[2,3] YU Xinyou;LI Tianzhi;MIAO Lizhong;XIE Jinwen;WANG Wenxiu(Shandong Lüdu Bio-Industry Co.,Ltd.,Binzhou,Shandong 256600;Shandong Binzhou Animal Science&Veterinary Medicine Academy,Binzhou,Shandong 256600;Shandong Academician Workstation,Binzhou,Shandong 256600)

机构地区：[1]山东绿都生物科技有限公司,山东滨州256600 [2]山东省滨州畜牧兽医研究院,山东滨州256600 [3]山东省院士工作站,山东滨州256600

出　　处：《中国家禽》2023年第12期47-52,共6页China Poultry

基　　金：滨州市农社领域科技创新政策引导计划项目(2022KPTY026);山东省外专双百计划项目(WST2018014);山东省现代农业产业技术体系家禽创新团队计划资助项目(SDAIT-11-16)。

摘　　要：为建立同时检测鸭甲型肝炎病毒3型(DHAV-3)和鸭圆环病毒(DuCV)的双重荧光定量PCR方法,试验根据NCBI中收录的DHAV-3 VP1和DuCV cap基因序列,分别设计了1对特异性引物和1条TaqMan探针,建立了同时检测DHAV-3和DuCV的双重荧光定量PCR,对该方法的特异性、敏感性和重复性进行评估,并采用该方法对临床样品进行检测。结果显示:建立的方法不与其他常见鸭病原发生交叉反应,对DHAV-3和DuCV的检测灵敏度分别为3.8 copies/μL和7.3 copies/μL,批内和批间变异系数均小于2%,从65份临床样品中分别检出DHAV-3和DuCV阳性样品11份和28份,与现有地方标准符合率分别为95.4%和90.8%。研究表明,建立的双重荧光定量PCR特异性强、敏感性高、重复性好,可用于DHAV-3和DuCV的快速检测。To establish a duplex real-time PCR assay for simultaneously detecting duck hepatitis virus type 3(DHAV-3)and duck circovirus(DuCV),according to the regions of VP1 gene of HAV-3 and cap gene of DuCV,two pairs of primers and twofluorescent probes were designed,the specificity,sensitivity and repeatability of the established duplex real-time PCR were evalu-ated,and the clinical samples were detected with the established assay.The results showed that the established assay did not crossreact with other common duck-origin pathogens,the detection limit of which was 3.8 copies/μL of DHAV-3 and 7.3 copies/μL of DuCV,respectively,the coefficient of variation within and between groups were all below 2%.There were 11 positive samples of DHAV-3 and 28 positive samples of DuCV detected from 65 clinical samples,respectively.The coincidence rates with existing local standards were 95.4%and 90.8%.In conclusion,the assay established in this study was highly specific,sensitive and re-peatable for rapid detection of DHAV-3 and DuCV.

关 键 词：鸭甲型肝炎病毒3型 鸭圆环病毒 双重荧光定量PCR 

分 类 号：S855.3[农业科学—临床兽医学]