猪繁殖与呼吸综合征病毒纯化方法

PRRSV(Porcine Reproductive and Respiratory Syndrome Virus,猪繁殖与呼吸综合征病毒) was propagated in MARC-145 cells and purified by sucrose density gradient separation.Polyethylene glycol 8000(Sigma Chemical Co.,St.Louis,MO) at a final concentration of 8%(wt/vol) was added to concentrate the virus.After incubation at 4°C overnight,the precipitated virus was centrifuged at 10,800xg for 20 min.The virus pellet was resuspended in TNE buffer(50 mM Tris-HCl,pH 7.5,100 mM NaCl and 1 mM EDTA).The resuspended virus was layered on 10 to 60%(wt/wt) sucrose gradients and centrifuged at 90,000xg at 4°C overnight.The purified virus band was collected, and the density(g/cm3) was determined. Centrifugation was performed at 90,000xg for 1h to pellet the purified virus, and the pellet was resuspended in TNE buffer.

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