6种猪病毒病微流控芯片多重检测方法的建立OA

In order to simultaneously detect the classical strain of porcine reproductive and respiratory syndrome virus(PRRSV),PRRSV highly pathogenic strain(HP-PRRSV),classical swine fever virus(CSFV),pseudorabies virus(PRV),porcine circovirus type 2(PCV2)and porcine parvovirus(PPV),primers were designed according to the genetically conserved regions of the 6 viruses,respectively,and embedded in reaction pools of isothermal amplification microfluidic chip,a microfluidic chip method was established to simultaneously detect all the above viruses through optimizing the concentration of primers,reaction temperature and time.The results showed that the lowest detection limits of the established method for the plasmid templates of the 6 viruses were all 103 copies/µL;specificity analysis showed that viruses or samples presented an"S"-shaped amplification curve only in the reaction pool corresponding to their specific detection indicators;good reproducibility was observed,the intra-and inter-batch coefficients of variation were less than 5%;for 40 clinical samples,the compliance rate of the method with commercial quantitative fluorescence detection kits reached above 95%.In conclusion,a microfluidic chip method was successfully established for 6 porcine viral diseases,which presented high sensitivity,strong specificity and good reproducibility,achieving joint detection for multiple animal diseases.