2.3 Virus isolation
Cerebrum specimens from the dead goat were handled as describe above, the supernatants were filtrated through a 0.22 um filter (Millipore, USA) and incubated with monolayer PK15 cells. The cells were cultured in Dulbecco modified eagle medium (DMEM) supplied with 5% newborn bovine serum (NBCS) and 1% streptomycin/penicillin. After one-hour incubation, the supernatants were removed and replaced with maintenance medium (DMEM with 2% NBCS and 1% streptomycin/penicillin). When obvious cytopathic effects (CPEs) were observed, the cells and supernatants were harvested. After three cycles of thawing and refreezing, PCR was performed to detect the presence of PRV nucleic acid in the supernatants. Isolated and identified PRV virus was designated as YNG strain.