人SUMO-2 shRNA干扰表达载体构建及其对细胞增殖的影响  

Construction of SUMO-2 shRNA knockdown vector and its effect on cell proliferation

作　　者：李向云[1] 黄宏[1] 邢伟[1] 郭韡[1] 何静[1] 孙志亚[1] 徐祥[1] 

机构地区：[1]第三军医大学附属大坪医院野战外科研究所(创伤烧伤与复合伤国家重点实验室第一研究室),重庆400042

出　　处：《中国现代医学杂志》2013年第28期1-5,共5页China Journal of Modern Medicine

基　　金：创伤烧伤与复合伤国家重点实验室基金(No:SKLZZ200907);国家自然科学基金(No:30970651)

摘　　要：目的构建干扰人SUMO-2表达的shRNA重组质粒载体,并筛选下调人SUMO-2表达的稳定细胞株。方法设计并合成针对人SUMO-2的shRNA寡核苷酸,并将其克隆至pLKO.1质粒载体上,转染HCT116细胞后经嘌呤霉素筛选获稳定表达细胞株,并通过免疫印迹鉴定SUMO-2的表达。结果成功构建干扰人SUMO-2表达的pLKO.1质粒载体,可在HCT116细胞中稳定表达,且能有效抑制SUMO-2蛋白质的表达。功能研究初步证明干扰SUMO-2表达可抑制细胞增殖并使处于细胞周期G1期的细胞比例上升。结论获得了干扰人SUMO-2表达的pLKO.1-shRNA质粒载体,以及下调SUMO-2表达的HCT116稳定细胞株。[Objective] To construct shRNA recombinant vector interfering human SUMO-2 and screen the stable cell line of SUMO-2 knockdown. [ Methods ] The shRNA oligos was designed and synthesized and subcloned into pLKO.1-TRC cloning vector. A stable cell line was generated by transfecting HCT116 cells and puromycin selec- tion. The expression of SUMO-2 was identified by Western blot. [Results] The pLKO.I-shRNA was successfully constructed, which was stably expressed in HCT116 cells and could interfere the expression of human SUMO-2 ef- fectively. Preliminary function study showed that SUMO-2 knockdown inhibited cell proliferation and decreased the percentage of S phase in cell cycle. [Conclusions] pLKO.I-shRNA interfering the expression of human SUMO-2 and an HCT116 stable cell line of SUMO-2 knockdown were obtained.

关 键 词：SHRNA干扰 SUMO-2 下调 稳定细胞株 细胞增殖 细胞周期 

分 类 号：Q291[生物学—细胞生物学]