1.Development and application of a triplex TaqMan fluorescent quantitative PCR assay for simultaneous detection of Senecavirus A,foot-and-mouth disease virus and porcine teschovirus
Shiqi GAN ; Qianhe WEI ; Yuchen NI ; Jianbo NI ; Xiuling ZHAO ; Wanyu DONG ; Yings-han ZHOU ; Xiaodu WANG
Chinese Journal of Veterinary Science 2025;45(1):22-29
Primers and probes were designed based on the conserved regions of Senecavirus A(SVA),foot-and-mouth disease virus(FMDV),and porcine teschovirus(PTV)and used to devel-op a TaqMan fluorescent quantitative PCR method for detecting the above-mentioned three viru-ses.The triplex fluorescent quantitative PCR system was developed using recombinant positive plasmids containing conserved sequences of the three viruses as templates.After optimizing the conditions,the specificity,sensitivity,repeatability,standard curve,and mixed infection model were evaluated,and the constructed triplex fluorescent quantitative PCR was used for clinical detection.The results showed that this method could specifically detect SVA,FMDV and PTV without cross-reactivity with other pathogens with the minimal detection concentrations for SVA,FMDV,and PTV as low as 1X101 copies/μL,respectively.The coefficients of variation within and between groups were less than 5%.Furthermore,none of the three viruses were detected in 126 samples.The above results indicate that this method is highly specific,sensitive,and stable,making it suit-able for clinical detection.
2.Development and application of a triplex TaqMan fluorescent quantitative PCR assay for simultaneous detection of Senecavirus A,foot-and-mouth disease virus and porcine teschovirus
Shiqi GAN ; Qianhe WEI ; Yuchen NI ; Jianbo NI ; Xiuling ZHAO ; Wanyu DONG ; Yings-han ZHOU ; Xiaodu WANG
Chinese Journal of Veterinary Science 2025;45(1):22-29
Primers and probes were designed based on the conserved regions of Senecavirus A(SVA),foot-and-mouth disease virus(FMDV),and porcine teschovirus(PTV)and used to devel-op a TaqMan fluorescent quantitative PCR method for detecting the above-mentioned three viru-ses.The triplex fluorescent quantitative PCR system was developed using recombinant positive plasmids containing conserved sequences of the three viruses as templates.After optimizing the conditions,the specificity,sensitivity,repeatability,standard curve,and mixed infection model were evaluated,and the constructed triplex fluorescent quantitative PCR was used for clinical detection.The results showed that this method could specifically detect SVA,FMDV and PTV without cross-reactivity with other pathogens with the minimal detection concentrations for SVA,FMDV,and PTV as low as 1X101 copies/μL,respectively.The coefficients of variation within and between groups were less than 5%.Furthermore,none of the three viruses were detected in 126 samples.The above results indicate that this method is highly specific,sensitive,and stable,making it suit-able for clinical detection.

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