RNAi对HCV ns5b基因表达的抑制作用

目的：研究RNAi(RNA interference)效应对HCV ns5b基因表达的抑制作用．方法：根据HCV ns5b基因序列及其二级结构特征，依据Tuschl等设计siRNA的原则，确定并合成了针对ns5b基因的siRNAs；利用电转化方法将合成的siRNAs转染NS5B-EGFP HepG2细胞株，以未转染的细胞为对照；细胞爬片后经DAPI染色，荧光显微镜下观察和半定量RT-PCR法检测siRNAs的抑制效应．结果：化学合成的siRNAs可以抑制HCV ns5b基因的表达，效率可达70％-80％以上，结论：在细胞水平上，化学合成的siRNAs可以抑制HCV ns5b基因的表达，为利用RNAi抑制HCV RNA复制的研究奠定基础。

Inhibitory effect of RNAi on HCV ns5b gene expression

AIM: To inhibit HCV ns5b gene expression with RNAi. METHODS: According to the gene sequence and secondary structure of HCV ns5b , we designed the siRNAs targeting ns5b gene following the requirement from Tuschl et al and it was synthesized in Dharmacon company. HepG2 cells stably expressing NS5B EGFP protein were transfected by synthesized siRNAs with electroportion, the non transfected cells and non specific siRNAs transfected cells were taken as controls. Inhibitory effect of siRNAs was detected under fluorescence microscope with DAPI dyeing and by semi quantitative RT PCR. RESULTS: Synthesized siRNA inhibited HCV ns5b gene expression by 70%-80%. CONCLUSION: At the cell level, inhibition of HCV ns5b gene expression by synthesized siRNA provides the new method for anti HCV study.