2.3 PCV2 detection with real-time PCR assay
PCV2 real-time PCR screening was performed as previously described (Wang et al., 2019). Briefly, PCR reactions were performed in a 20-ul reaction containing 4 ul of extracted DNA, 0.2 uM each of forward and reverse primers, 0.1 uM each of probes, and 10 ul of Luna® Universal Probe qPCR master mix (NEB, MA, USA). PCV2 was detected using Quantstudio5 real-time system (Appiled Biosystems, USA). The PCR condition consisted of initial denaturation at 94°C for 30s followed by 45 cycles of 94°C for 15s, and 60°C for 45s.