人源性14-3-3σ真核表达载体的构建及鉴定  

作　　者：甘露[1] 朱阅伟[2] 周美娟[1] 欧程山[1] 丁振华[1] 

机构地区：[1]南方医科大学公共卫生与热带医学学院放射医学系 [2]南方医科大学

出　　处：《科技信息》2010年第22期24-25,共2页Science & Technology Information

基　　金：广东省自然科学基金博士启动项目资助(No.9451051501002535);南方医科大学2008-2009学年学生课外科研学术活动立项项目资助(No.2008kw041);南方医科大学公共卫生与热带医学学院院长基金(2009年)

摘　　要：目的:构建人源性14-3-3σ真核表达载体pCMV-Myc-SFN,并观察其在真核细胞中的表达。方法:通过反转录-聚合酶链反应从HeLa细胞(人宫颈癌细胞系)中获得编码14-3-3σ的cDNA,定向克隆至真核表达载体pCMV-Myc中,经酶切和测序鉴定正确后,应用脂质体法转染HeLa细胞,并通过蛋白免疫印迹法检测细胞内14-3-3σ的表达。结果:构建了真核表达载体pCMV-Myc-SFN,将其转染HeLa细胞48h后,蛋白免疫印迹法检测到细胞内14-3-3σ的表达显著增高。结论:成功构建了真核表达载体pCMV-Myc-SFN,为研究14-3-3σ在上皮细胞源性肿瘤中的作用奠定了基础。Objective：To construct the eukaryotic expression vector of human 14-3-3σ, pCMV-Myc-SFN, and to examine its expression in vitro. Methods：Total RNA was extracted from the HeLa cells and reverse transcription-polymerase chain reaction （RT-PCR） was per-formed to obtain the cDNA fragment encoding 14-3-3σ, which was inserted into the pCMV-Myc vector. And the new construct, pCMV-Myc-SFN was confirmed by restriction enzyme digestion and DNA sequencing. HeLa cells were transfected with the pCMV-Myc-SFN vector and pCMV-Myc vector, respectively. The expression of 14-3-3σ was detected by Western blotting. Results： The eukaryotic expression vector pCMV-Myc-SFN was constructed, and significant increase of 14-3-3σ expression was detected in the HeLa cells 48 hours after transfection. Conclusion：The eukaryotic expression vector pCMV-Myc-SFN was constructed successfully.

关 键 词：14-3-3σ 上皮性恶性肿瘤 真核表达载体 

分 类 号：Q782[生物学—分子生物学]