Development of a duplex TaqMan real-time PCR assay for simultaneous detection and differentiation of canine coronavirus and canine parvovirus
A dual TaqMan fluorescence quantitative PCR assay for detection of canine coronavirus and canine parvovirus was established in this study.Primers and probes were designed for 3'UTR of CCoV and VP2 of CPV.After the reaction system and conditions were optimized,the sensitivity,specificity and repeatability of the method were verified and clinical samples were tested.The results showed that the standard curves R2 of CCoV and CPV positive reference plasmids ranged from 1 ×107 copies/μL to 1×102 copies/μL,and the linear relationship was good.The minimum detection limits of CCoV and CPV were both 5 copies/μL.There was no cross-reaction with other pathogens,and the co-efficient of variation within and between groups was less than 1.38％.The results indicated that this method had high sensitivity,strong spe-cificity and good repeatability.Compared with ordinary PCR,the detection rate of CCoV and CPV using this method was 37.2％higher and 40.7％higher.Therefore,the PCR method established in this study had a good effect and could provide help for clinical diagnosis and epide-miological investigation.
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维普
