猪传染性胃肠炎病毒、猪流行性腹泻病毒和猪轮状病毒三重PCR检测方法的建立

摘要

通过设计三对引物分别扩增猪传染性胃肠炎病毒（TGEV）、猪流行性腹泻病毒（PEDV）及猪轮状病毒（RV）的特异性基因,建立了一个能对TGEV、PEDV及RV感染鉴别诊断的三重PCR方法。该方法能同时检测TGEV的M基因252bp片段、PEDV的N基因540bp片段及RV的VP7基因412bp片段,而对CSFV、PCV2、PRRSV、PRV等无扩增,其检测TGEV、PEDV及RV的极限为100pg核酸/μL;用该方法对临床收集的26份粪便样品进行检测,结果表明：建立的多重PCR方法具有特异性强,强灵敏度高等特点,能用于临床诊断及流行病学调查。%Three pairs of primers amplified genes of transmissible gastroenteritis virus（TGEV）,porcine epidemic diarrhea virus（PEDV） and porcine rotavirus（RV） respectively were designed to develop a method of differential diagnosis of TGEV,PEDV and RV.The established PCR can detect M gene with the length of 252bp of TGEV,N gene with the length of 540 bp of PEDV and VP7 gene with the length of 412 bp of RV.Negative results when using CSFV,PCV2,PRRSV and PRV as control were obtained.The detection limit of this method was 100-pg nucleotide per PCR reaction.Twenty six diarrheal fecal samples collected from viral diarrheal piglet were submitted to detect TGEV,PEDV and RV,and the result showed that this multiplex RT-PCR method with the characteristics of high sensitivity and high specificity can be widely used in clinical diagnosis and epidemiological investigation.