Porcine pseudorabies virus (PRV) variant PRV-ZJ01 and application thereof
A technology of PRV-ZJ01 and porcine pseudorabies, applied in antiviral agents, viruses/phages, medical preparations containing active ingredients, etc., to achieve good immunogenicity and safety
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[0032] 1. Virus isolation and PCR identification
[0033] The collected brain tissue of diseased pigs was shredded and mixed (the weight-to-volume ratio of tissue to sterilized PBS was 1:10), centrifuged at 12,000 rpm for 10 min, and the supernatant was filtered through a 0.22 μm filter for virus isolation. Inoculate 1mL of the filtered supernatant of the disease material into BHK-21 cells that have just grown into a single layer, incubate at 37°C for 1h, discard the liquid in the culture flask, and add 2% DMEM in a solution containing 5% CO 2 Cells were cultured in a 37°C cell culture incubator, and the cells were observed for lesions every day. After 70% of the cells had lesions, they were frozen and thawed three times, and BHK21 cells were inoculated. Virus plaques were purified 3 times, and the gE gene was amplified by PRV PCR method. Reaction system: 1 U AccuPrime TM pfx DNA Polymerase with 2.5μL 10×AccuPrime TM pfx Mix (Invitrogen), upstream and downstream primers...
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