猪圆环病毒3型和4型双重PCR检测方法的建立  

Development of a duplex PCR for detecting porcine circovirus types 3 and 4

作　　者：侯承尧 徐通 崔建涛[1] 田润博 郑兰兰[1] 赵丽[2] 陈红英[1] HOU Chengyao;XU Tong;CUI Jiantao;TIAN Runbo;ZHENG Lanlan;ZHAO Li;CHEN Hongying(College of Veterinary Medicine,Henan Agricultural University,Zhengzhou 450002,China;College of Veterinary Medicine,Henan University of Animal Husbandry and Economy Zhengzhou 450046,China)

机构地区：[1]河南农业大学动物医学院,河南郑州450002 [2]河南牧业经济学院动物医药学院,河南郑州450046

出　　处：《中国兽医学报》2021年第9期1682-1686,1703,共6页Chinese Journal of Veterinary Science

基　　金：中原千人计划资助项目(204200510015);河南省高校科技创新团队支持计划资助项目(19IRTSTHN007);河南牧业经济学院科技创新团队资助项目(2018KYTD18)。

摘　　要：为了检测和鉴别猪圆环病毒3型(PCV3)和4型(PCV4),根据GenBank已登录的PCV3和PCV4基因序列,在PCV3 Rep和PCV4 Cap基因的高度保守区域设计并合成2对特异性引物,经过双重PCR反应条件优化,建立了能够同时检测PCV3/4的双重PCR方法。结果显示,该方法能同时扩增出138(PCV3)和391 bp(PCV4)特异片段,而对PCV2、猪伪狂犬病病毒、猪细小病毒、猪繁殖与呼吸综合征病毒、猪瘟病毒和猪流行性腹泻病毒核酸扩增结果均为阴性。PCV3/4的检测极限分别为74.5,90.2拷贝/μL。经临床样本检测结果表明,本试验建立了同时检测PCV3/4双重PCR方法,且具有准确、可靠、灵敏度高、特异性强的特点,为PCV3/4的临床检测和鉴别诊断提供了技术支持。To detect and differentiate porcine circovirus type 3(PCV3)and porcine circovirus type 4(PCV4)simultaneously,two pairs of specific primers were designed and synthesized based on the highly conserved regions of the Rep gene of PCV3 and the Cap gene of PCV4 according to the published genome sequences of PCV3 and PCV4 in GenBank.After optimizing the reaction conditions,a duplex PCR method was developed to detect PCV3/4 simultaneously.The results indicated that the 138 bp specific fragment of PCV3 and 391 bp specific fragment of PCV4 were simultaneously amplified,but the amplification results of porcine circovirus type 2,porcine pseudorabies virus,porcine parvovirus,porcine reproductive and respiratory syndrome virus,porcine classical fever virus,and porcine epidemic diarrhea virus were negative.The limits of detection of the assay were 74.5 copies/μL for PCV3 and 90.2 copies/μL for PCV4.The detection results of clinical samples showed that the duplex PCR method established in this study was accurate and reliable,with high sensitivity and good specificity,which can provide technical support for the differential diagnosis and prevention and control of PCV3 and PCV4.

关 键 词：猪圆环病毒3型 猪圆环病毒4型 Rep基因 Cap基因 双重PCR 

分 类 号：S852.65[农业科学—基础兽医学]