猪流行性腹泻病毒RT-PCR检测方法建立及初步应用研究

Development and Preliminary Application of a RT-PCR for Detection of Porcine Epidemic Diarrhea Virus

为建立一种特异、敏感的PEDV的检测方法。根据GenBank中公布的猪流行性腹泻病毒CV777株的M基因序列,设计了一对特异性引物,扩增长度为680bp的片段。将PCR产物进行测序,与CV777株、SH5株M基因的同源性分别为99.2%和97.6%。试验表明:该方法具有较好的特异性和敏感性,并能检测到PEDV的RNA浓度下限为100pg/μL。从福建省的3个地区共采集58份哺乳仔猪腹泻样品,并应用此方法进行检测,其阳性检出率为87.9%。

In order to establish specific and sensitive detection methods of porcine epidemic diarrhea virus（PEDV）.A pair of primer was designed to amplify a product about 680 bp,according to the sequence of the M gene sequence of PEDV-CV777 strain of PEDV in Genbank.The amplified product was sequenced.Then the homology comparisons among the obtained sequence and the M Gene of the CV777 and SH5 stains were performed.The results showed that it shared 99.2% and 97.6% homology with M Gene of the CV777 and SH5,respectively.This method could be used to test PEDV,and the minimum detectable concentration of the PEDV RNA was 100 pg/μL.The method has been used to detect 58 portion specimens of diarrheal swine from three areas named A,B,and C in Fujian Province,and the positive tested rate of PEDV was 87.9%.