携带Oct4、Klf4、Sox2、c-myc和EGFP基因小鼠肿瘤细胞株的建立  

Construction of mouse tumor cell lines harboring Oct4, Klf4, Sox2, c-myc and EGFP genes

作　　者：贾琦珺[1] 史俊文[1] 申洪芬[1] 贾俊双[1] 肖东[1,2] 姚开泰[1] 

机构地区：[1]南方医科大学肿瘤研究所,广东广州510515 [2]南方医科大学比较医学研究所暨实验动物中心,广东广州510515

出　　处：《热带医学杂志》2012年第4期366-368,F0002,共4页Journal of Tropical Medicine

基　　金：国家自然科学基金委员会-广东省联合基金重点项目(u0732006);国家自然科学基金(30271177);广东省自然科学基金(021903和9151063101000015);广东省卫生厅基金(A2007359);南方医科大学优秀中青年科技人才库科研资助金;广州地区科学仪器协作共用网专用基金(2006176)

摘　　要：目的构建稳定携带Oct4、Klf4、Sox2、c-myc(以下简称为OKSM)和增强型绿色荧光蛋白(enhanced green fluorescent protein,EGFP)基因的小鼠肿瘤细胞株。方法将载体pLentG-KOSM用NotI、XbaI线性化,并进行聚合酶链式反应(polymerase chain reaction,PCR),以鉴定其结构。按Invitrogen公司推荐的标准程序进行慢病毒包装和确认慢病毒是否成功生产;携带OKSM和EGFP基因的慢病毒感染小鼠肝癌细胞(Hep1-6)、小鼠乳腺癌细胞(4T1)和小鼠肺癌细胞(LLC),以建立相应病毒感染体系。结果 pLentG-KOSM确实含有OKSM4种基因,按标准程序生产的携带OKSM和EGFP基因的慢病毒上清高效率感染Hep1-6、4T1及LLC,成功建立稳定携带OKSM和EGFP基因的小鼠肿瘤细胞株。结论成功构建携带OKSM和EGFP基因的Hep1-6、4T1、LLC细胞株,为相关的后续研究打下了良好基础。Objective To generate mouse tumor cell lines harboring Oct4, Klf4, Sox2, c-myc （OKSM） and enhanced green fluorescent protein （EGFP） genes. Methods Recombinant lentiviral vector of pLentG-KOSM was constructed and indetified by enzyme digestion with Not I,Xba I and polymerase chain reaction （PCR）. According to the standard protocol from Invitrogen,lentiviruses were produced, followed by confirming that lentiviruses were successfully made through EGFP assay under fluorescent microscope after infecting 293FT cells. Lentiviruses harboring OKSM and EGFP genes were employed to infect mouse tumor cells[i.e.,mouse hepatocellular carcinoma cells（Hep1-6）,mouse breast cancer cells（4T1）and mouse lung cancer cells（LLC）]. Results Lentiviral vector pLentG-KOSM was confirmed to be correct by enzyme digestion and PCR.Lentivirus supernatant harboring OKSM and EGFP genes efficiently infected Hep1-6,4T1 and LLC. Conclusion The mouse tumor cell lines harboring OKSM and EGFP genes were successfully constructed,which will lay a solid foundation for further research.

关 键 词：OCT4 KLF4 SOX2 C-MYC 肿瘤细胞重编程 

分 类 号：R34[医药卫生—基础医学]