猪δ冠状病毒荧光定量RT-PCR检测方法的建立  

Establishment of fluorescence quantitative RT-PCR for detection of porcine deltacoronavirus

作　　者：孙彦婷[1] 康宇 路亚男 井汇源 董望 李向辉[1] 王金合[1] 唐光武[1] SUN Yanting;KANG Yv;LU Yanan;JING Huiyuan;DONG Wang;LI Xianghui;WANG Jinhe;TANG Guangwu(Henan University of Animal Husbandry and Economy,Zhengzhou Henan 450046)

机构地区：[1]河南牧业经济学院,河南郑州450046

出　　处：《现代牧业》2023年第3期12-17,共6页Modern Animal Husbandry

基　　金：河南省重点研发与推广专项项目(202102110247);河南省教育厅高等学校重点科研计划项目(16A230007);河南牧业经济学院博士启动基金项目(53000212);河南牧业经济学院预防兽医学重点学科建设项目(MXK2016102)。

摘　　要：建立一种快速检测猪δ冠状病毒(PDCoV)的方法。根据GeneBank收录的PDCoV M基因序列,设计两对特异性引物和TaqMan探针,对PDCoV TaqMan荧光定量RT-PCR反应条件进行优化。Ct值与质粒模板之间呈现良好的线性关系,标准方程为y=-3.6421x+34.194,相关系数R 2=0.9992。荧光定量RT-PCR方法仅对PDCoV呈现特异性扩增,无交叉反应。敏感度试验结果显示,最低检出量为1.25×10^(1)copies/μL,灵敏度高。重复性试验结果显示,批内和批间测定均具有良好的重复性。TaqMan荧光定量RT-PCR检测方法是一种可靠的快速检测PDCoV的方法。The purpose of the experiment was to establish a rapid testing method for porcine deltacoronavirus(PDCoV).Based on the PDCoV M gene sequence included in GeneBank,two pairs of specific primers and TaqMan probes were designed to optimize the reaction conditions of PDCoV TaqMan fluorescence quantitative RT-PCR.There was a good linear relationship between Ct value and plasmid template,and the standard equation was y=-3.6421x+34.194,and correlation coefficient was R 2=0.9992.The method only showed specific amplification of PDCoV without cross-reaction.The sensitivity test showed that the minimum detection amount was 1.25×10^(1)copies/μL with high sensitivity.The results of repeatability test showed that both within and between batches had good repeatability.The TaqMan fluorescence quantitative RT-PCR detection method is a reliable and rapid method for diagnosing PDCoV.

关 键 词：猪δ冠状病毒 M基因 荧光定量RT-PCR TAQMAN 

分 类 号：S855.3[农业科学—临床兽医学]