Establishment and application of TaqMan fluorescent probe- based quantitative PCR assay for detection of Porcine circovirus type 2

To detect quickly and accurately detect Porcine circovirus type 2( PCV-2) and quantify the number of viral copies,a pair of specific primers and Taq Man probes were designed according to the conservative sequence of PCV-2 in Genbank( accession number: FJ644559.1).The standards were prepared,and then the fluorescent quantitative PCR assay for detection of PCV-2 was established and used to detect the clinical samples.The results showed that the assay had good linearity within the template ranges from 1 × 101 copies/μL to 1 ×108copies/μL,the correlation coefficient was up to 0.999,and its sensitivity was 100 times that of routine PCR assay; the assay was negative with Porcine parvovirus( PPV),Porcine pseudorabies virus( PRV),Calssical swine fever virus( CSFV) and Porcine reproductive respiratory syndrome virus( PRRSV),there was no cross-reaction.The intra and inter-assay coefficients of variation were less than 2.50%.The positive detection rate of PCV-2 using the fluorescent quantitative PCR assay was 38% higher than that using conventional PCR method in the clinical detection.The result indicates that a Taqman fluorescent probe-based quantitative PCR assay is successfully established which can be used for the clinical detection of PCV-2 infection and quantification of the number of PCV-2 copies.