利用RNA干扰机制抑制猪繁殖与呼吸综合征病毒的增殖.pdfVIP

第 21 卷 3 期 中 国 病 毒 学 21 （3 ）：226-230 2006 年 5 月 VIROLOGICA SINICA May 2006 利用RNA 干扰机制抑制猪繁殖与呼吸综合征病毒的增殖 高晓飞，包晶晶，陈勇军，陈溥言** （南京农业大学动物医学院，江苏南京210095 ） RNA Interference Inhibits Replication of Porcine Reproductive and Respiratory Syndrome Virus GAO Xiao-fei，BAO Jing-jing ，CHEN Yong-jun，CHEN Pu-yan** (College of Animal Veterinary Medicine，Nanjing Agricultural University, Nanjing 210095, China) Abstract ：RNA interference (RNAi) was performed to study the inhibition of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) replication. Two short interfering RNAs (siRNA) specific for N gene of PRRSV were designed using an online software. The siRNA was shown to inhibit PRRSV replication in cell lines. Two siRNA from PRRSV N gene expressed by vectors were shown to have a clear RNAi effect on PRRSV replication in MARC-145 cells. TCID50 of progeny virus at different time, the occurrence time of CPE and IFA results in each group gave the evidence that the antiviral effects could extend to 120h. This report successfully revealed RNAi phenomenon of PRRSV replication procedure in MARC-145 cells and showed that vector-based RNAi technology would give a new vaccine design in PRRSV therapy. Key words: Porcine reproductive and respiratory syndrome (PRRS); TCID50; N gene; CPE ； 摘要：本文探讨依靠RNAi技术对猪繁殖与呼吸综合征病毒( PRRSV) 增殖的干扰作用。筛选到针对编码PRRS病毒 核衣壳蛋白的N基因的两处靶序列作为候选片段，在MARC-145细胞上进行基因干扰实验研究。成功观测到由载 体表达的小干扰RNA (siRNA)在MARC-145细胞中对PRRS病毒增殖的抑制现象。通过选取不同时间段对病毒进行 TCID50检测，以及对CPE 出现时间进行观察和免疫荧光技术，得到RNA干扰对PRRS病毒增殖抑制作用的动态数据。 证实在真核细胞水平上，RNA干扰机制可以抑制PRRS病毒的增殖。实验结果表明，依靠载体表达的RNA干扰技