Abstract Several genome sequences of Foot-and-mouth disease virus (FMDV) were obtained from GenBank. After multiplex alignment, three sets of primers and probes were designed according to 3D, VP3 and 3C region of FMDV. Three sets of primers and probes were used to establish FMDV species-specificity, O type-specificity and AsiaⅠtype-specificity Real-time RT-PCR, respectively. Standard S shape curves occurred after Real-time RT-PCR amplification on Lightcycler machine. All three newly established methods had good specification for they could differentiate FMDV from Infectious bovine rhinotracheitis virus, Swine transmissible gastroenteritis virus, Akabane virus and Porcine respiratory corona virus. FMDV species-specificity Real-time RT-PCR could successfully amplify O type(O1 strain and O2 strain) and AsiaⅠtype (AsiaⅠ1 strain and AsiaⅠ2 strain) FMDV. O type-specificity Real-time RT-PCR could successfully amplify O type(O1 strain and O2 strain) FMDV but failed to amplify AsiaⅠ (AsiaⅠ1 strain and AsiaⅠ2 strain) FMDV. AsiaⅠtype-specificity Real-time RT-PCR could successfully amplify AsiaⅠ(AsiaⅠ1 strain and AsiaⅠ2 strain) FMDV but failed to amplify O type (O1 strain and O2 strain) FMDV. All methods could detect 10 copies of DNA template, closely reach the detection limit. The whole Real-time RT-PCR reaction could be finished in 70 min. These methods have potential to apply in FMDV screen and differential detection in laboratory.
