中文摘要：

目的构建转录因子NF-κB的RNA干扰（RNAi）质粒,为研究NF-κB参与的细胞信号通路及以其为靶点的基因治疗提供稳定转染的RNAi质粒。方法设计针对NF-κB的shRNA特异性序列,应用基因重组技术克隆到pSilencer 1.0-U6表达载体,EcoRⅠ和ApaⅠ双酶切及DNA测序鉴定重组克隆;脂质体转染RNAi重组载体至前列腺癌细胞PC-3后,Western blot分析各组NF-κB蛋白表达水平。结果双酶切和DNA测序证实shRNA正确插入pSilencer 1.0-U6质粒;Western blot结果显示与空质粒对照组比较NF-κB转染组细胞NF-κB表达明显下调。结论成功构建人NF-κB基因RNAi质粒,为研究NF-κB参与的细胞信号通路提供了稳定转染细胞的干扰质粒,为靶向NF-κB的基因治疗提供了有力工具。

英文摘要：

Objective To construct RNA interference plasmid targeting NF-κB,it provided stable RNA interference plasmid for NF-κB study involved in cell signaling pathway and NF-κB-targeting gene therapy.Methods NF-κB-targeting shRNA sequences were designed and cloned into the expression vector pSilencer 1.0-U6.The recombinants were identified by using EcoRⅠ ＋ ApaⅠ double digestion and DNA sequencing,then transfected into prostate cancer cells PC-3 with Lipofectamine 2000.NF-κB protein expressions were detected by Western blot.Results shRNA was inserted into plasmid pSilencer 1.0-U6 correctly confirmed by double digestion and DNA sequencing.NF-κB expression was decreased in NF-κB transfected group compared to control group by the results of Western blot.Conclusion RNA interference plasmid targeting NF-κB is successfully constructed,it provides stable RNA interference plasmid for the research of NF-κB participating in cell signaling pathway,and offers favorable tool for NF-κB-targeting gene therapy.