Abstract: In order to construct real-time quantitative PCR assay for detection of African swine fever virus, this study was based on 23 isolates of gene sequence which encodes ASFV structural protein p72 in GenBank, then designing primers and probe. The reaction conditions were optimized by using different annealing temperature, different Mg^2＋ concentrations, different primers and probe concentrations. The real-time PCR system could automatically generate standard curve, testing repeatability, sensitivity and specificity. Wild boar samples were detected by this assay. The results showed optimal annealing temperature was 60 ℃, optimal Mg^2＋ concentration was 4 mmol/L, optimal primers and probe concentration were 0.8 and 0.3 μmol/L.The coefficients of variation of repeatability test were less than 1.3%, sensitivity tests could detect 10 copies/μL plasmids, the specificity was tested by detecting five others swine viruses and ASFV plasmid, only detection of ASFV plasmid appears amplification curve. In conclusion, constructed real-time quantitative PCR assay was rapid,sensitive and specific assay for detection of ASFV.

Key words: African swine fever; ASFV; real-time quantitative PCR