Detection of porcine parvovirus by quantitative real-time PCR assay

According to genome sequences of porcine parvovirus(PPV)published in GenBank,A pair of primers and a TaqMan probe were designed.The expected fragment was amplified from DNA of PPV-infected PK-15 cells.The purified PCR product was connected with pGEM-T-easy vector and then transferred into JM109.The standard recombinant plasmid was gained from positive bacterium clone.The plasmid PCR and plasmid sequence mensuration showed that the expected fragment was successfully cloned.104～108copies/μL standard recombinant plasmid DNA specimen were amplified by real-time quantitative PCR,which indicate that there is a good linear function in statistics between the Ct value and the concentration gradient of standard plasmid DNA specimen.Analysis of the dynamic curve showed that under this condition of the reaction,the sensitive degree is 102 copies.The construction of real-time quantitative PCR provided the basis for the early diagnosis of PPV infection.