慢病毒介导的RNAi靶向NDV P基因抑制其在CEF上的复制

Inhibition of Newcastle disease virus replication by lentivirus-mediated RNA interference targeting the P gene in chicken embryo fibroblasts

通过研究慢病毒介导的RNA干扰靶向新城疫病毒P基因抑制其在鸡胚成纤维细胞上的复制,从而为新城疫病毒的抗病毒研究奠定基础。①针对NDV P基因设计siRNA干扰序列,构建慢病毒(1entivirus)介导的RNAi重组载体;②携带有siRNA表达元件重组慢病毒包装及其滴度测定;③包装后重组慢病毒接种CEF细胞并接毒NDV,48 h分别进行Real time RT-PCR和NDV滴度测定,并观察细胞病变情况。结果表明,本研究针对NDVNA-1株P基因2个RNAi靶位(位于开放阅读框的641和827位)设计的RNAi-641和RNAi-827,对病毒复制具很强的抑制效果,且641位的重要性大于827位。

Abstract ： To lay the fdundation for the research of NDV anti-virus through the studies on inhibition of Newcastle disease virus replication by lentivirus mediated RNA interference targeting the P gene in CEF. Design a siRNA interference sequence of NDV P gene,and then build a RNAi recombinant vector mediated by lentivirus. The packaging and titer determination of the reorganized lentivirus which carried the siRNA expression elements. Firstly, using the packaged recombinant lentivirus vaccinate CEF cells as well as inoculate NDV on it; secondly, doing Real-time RTPCR and NDV titer determination at 48 h; lastly, observing the cell lesions. This study designed two RNAi target sites for the NDV NA-1 P gene （RNAi 641 and RNAi-827）,and the experimental results showed that both of them have strong inhibitory effects on viral replication.