中国兽医科学2024，Vol.54Issue(11)：1451-1457,7.DOI:10.16656/j.issn.1673-4696.2024.0178
猪流行性腹泻病毒通用实时荧光定量RT-PCR方法的建立与初步应用
Development and application of a universal fluorescent quantitative RT-PCR for detection of porcine epidemic diarrhea virus
摘要
Abstract
Our study introduces a fluorescent quantitative RT-PCR assay targeting the conserved M gene of porcine epidemic diarrhea virus(PEDV),with the sequences of primers and probes sharing 100％similarities with 96.93％of PEDV strains available including the sequences from GenBank and the ones in local databases.After optimizing the reaction conditions,the standard curve was established,and the correlation coefficient(R2)was 0.999 5,indicating a good linear relationship.There was no cross-reaction with porcine reproductive and respiratory syndrome virus,porcine rotavirus,Streptococcus suis,porcine circovirus,porcine parvovirus,pseudorabies virus,transmissible gastroenteritis virus of pigs,porcine deltacoronavirus,indicating a good specificity.The limit of detection was determined to be 2.10 copies/μL,with both intra-and inter-batch variations maintaining a coefficient of vari-ation under 2％.Comparative analysis of 162 clinical samples against quarantine protocols revealed a 100％positive coincidence rate,underscoring the assay's high sensitivity,specificity,and repro-ducibility.This advanced fluorescent quantitative RT-PCR method not only effectively identifies both G Ⅰ and GⅡ genogroups of PEDV but also significantly contributes to the early detection and control of the disease,offering a potent tool in virus identification and disease prevention.关键词
猪流行性腹泻病毒/M基因/实时荧光定量RT-PCR/TaqMan探针Key words
porcine epidemic diarrhea virus/Mgene/fluorescent quantitative RT-PCR/TaqMan probe分类
农业科技引用本文复制引用
李玮卓,骆焕东,林青青,刘萍,李军蓝,李静,李娟,成晶,陈振海,陈蕾..猪流行性腹泻病毒通用实时荧光定量RT-PCR方法的建立与初步应用[J].中国兽医科学,2024,54(11):1451-1457,7.基金项目
国家自然科学基金项目(32102680) （32102680）
江苏省科学技术厅-基础研究计划(自然科学基金)项目(BK20200931) （自然科学基金）
