中文摘要：

目的构建靶向CIP2A的shRNA真核表达质粒并探讨其对胃癌细胞增殖的调节作用。方法根据siRNA原理设计4对靶向CIP2A的siRNA序列,克隆到真核表达载体p GPU6/GFP/Neo中（shRNA-1、2、3和4）。脂质体转染人胃癌细胞系BGC-823,real-time PCR和Western blot法检测并筛选最佳抑制效率的shRNA表达质粒,CCK-8法检测对胃癌细胞增殖的影响。结果 4个靶向CIP2A的shRNA真核表达质粒经限制性酶切和测序证实基因已正确插入,转染效率可达到70%以上。转染后24、48和72 h,4个质粒均明显降低BGC-823细胞内CIP2A mRNA和蛋白的表达。相较于其他3个重组质粒,shRNA-1的抑制作用更为显著,且对胃癌细胞增殖有明显的抑制作用（P〈0.05）。结论成功构建的靶向CIP2A的shRNA真核表达质粒,可有效抑制胃癌细胞CIP2A的mRNA和蛋白表达,并抑制胃癌细胞的增殖,为今后研究CIP2A在胃癌中的作用机制奠定了基础。

英文摘要：

Objective To construct eukaryotic vectors expressing short hairpin RNAs（ shRNAs） targeting at the CIP2 A gene and to explore its effects on gastric cell line BGC-823. Methods Four oligonucleotides targeting the CIP2 A gene were synthesized and cloned into the eukaryotic expression plasmid p GPU6. The recombinant plasmids,p GPU6/GFP/Neo-CIP2A-shRNA-1,2,3 and 4,were introduced into BGC-823 cells by lipofectamine-mediated transfection and the infection rate was observed by fluorescence microscope. The gene silencing efficiency was measured by real-time PCR and Western blot. The effects on proliferation of BGC-823 cells were detected by CCK-8. Results DNA sequencing and enzyme digestion analysis confirmed the identity of the four recombinant shRNA expression vectors. Immunofluorescsence demonstrated that transfection efficiency was above 70%. Transfection of shRNA-1,2,3 and 4,significantly knocked down the expression of CIP2 A mRNA and protein at 24,48 and 72 h after transfection. Compared with the 2,3 and 4,shRNA-1 had the more strong inhibitory effect on the expression of CIP2 A mRNA and protein. The CCK-8 assay showed that the anti-proliferation effect on BGC-823 cells was significant（ P〈0. 05）. Conclusions The recombinant vector may effectively inhibit the expression ofCIP2 A in BGC-823 cells and depress the proliferation of BGC-823 cells.