中文摘要：

目的设计并构建靶向HBV的外源性microRNA表达载体,观察其对HBV表达和复制的抑制作用。方法以HBVP基因和X基因为靶基因,根据pcDNA6.2-G/W-miR载体要求,设计靶向HBV的microRNA序列,进行直接合成获得目的片段并克隆到载体中,测序正确的载体通过脂质体方法转染HepG2.2.15细胞,RT-PCR、乙肝五项定量和荧光定量PCR检测其对HBVmRNA和HBVDNA的抑制作用。结果质粒的转染效率为50%～60%,在这种状态下,外源性microRNA表达载体在HBVmRNA和HBsAg、HBeAg和HBVDNA水平上均能显著抑制HepG2.2.15细胞中HBV的复制和表达（P〈0.05）。结论外源性microRNA表达载体构建成功,并能抑制HBV靶基因的表达,为其在慢性HBV感染基因治疗中的应用奠定了基础。

英文摘要：

Objective To design and construct the artificial miRNA（amiRNA） vectors which targeted HBV,observe whether amiRNA-mediated RNA interference could decrease the replication level of HBV.Methods The amiRNA expression vectors targeted by HBV P and X gene were construted according to the pcDNA6.2-G/W-miR vector instruction.The plasmids were transient transfected by lipofectemine 2000 reagment.Semi-quantitative RT-PCR was utilized to observe suppression level of HBV mRNA;The MEIA was carried out to detect quantitatively HBsAg and HBeAg,the fluorescent quantitation PCR was utilized to observe the changes of HBV DNA copies.Results The transfection efficiency were 50%~60% by ECM analysis.Under such circumstances,the assay results showed that the four plasmids had obvious inhibited HBV replication and expression in HepG2.2.15 cell（P〈0.05）.Conclusion Artificial microRNA-mediated RNA interference can be high effectively block HBV expression in specific manners.Artificial microRNA targeted HBV may be a promising therapy approach for chronic HBV infection.