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.