中国兽医科学2017，Vol.47Issue(3)：334-340,7.DOI:10.16656/j.issn.1673-4696.2017.03.010
猪德尔塔冠状病毒RT-PCR检测方法的建立及应用
Establishment and application of RT-PCR method for detection of porcine deltacoronavirus
摘要
Abstract
This experiment was conducted to develop a sensitive and rapid detection method for porcine deltacoronavirus(PDCoV).A reverse transcription polymerase chain reaction(RT-PCR) for detecting PDCoV was established using a pair of specific primers based on the M gene (359 bp fragment) of PDCoV published in GenBank.Through the optimization of reaction components in the RT-PCR system,a sensitive and specific PDCoV RT-PCR assay was established,and its sensitivity,specificity and repeatability were tested.The results showed only PDCoV was successfully amplified with a target fragment of 359 bp in size,but the amplification results were negative for porcine reproductive and respiratory syndrome virus,classical swine fever virus,porcine epidemic diarrhea virus,transmissible gastroenteritis virus of swine,rabies virus,bovine viral diarrhea virus,porcine parvovirus,porcine circovirus type 2,pseudorabies virus,Streptococcus suis type 2 and Escherichia coli.Testing of the sensitivity of RT-PCR indicated that the lowest detection concentration was 1 ng/μL.The method was used to detect 110 clinically suspected cases of swine diarrhea,and the results showed the positive rate of PDCoV was 11.8％.Blast analysis showed homology between positive samples and other PDCoV nucleotide sequence was 92.2％ to 100％.Phylogenetic analysis based on M genes indicated that PDCoV strains had closer relationship with Chinese PDCoV strains than the strains isolated from other countries.The above result suggested that we successfully established an RT-PCR method for detection of PDCoV.The method can be used to monitor and epidemiologically investigate PDCoV infection.关键词
猪德尔塔冠状病毒/RT-PCR/检测Key words
porcine deltacoronavirus/RT-PCR/detection分类
农业科技引用本文复制引用
郑丽,王利丽,路超,杨春蕾,李富强,张莉,鄢明华..猪德尔塔冠状病毒RT-PCR检测方法的建立及应用[J].中国兽医科学,2017,47(3):334-340,7.基金项目
天津市农业科技转化与推广项目(201301030) （201301030）
