Smyd1干扰真核表达质粒及其稳定转染H9c2细胞系的构建  

Construction of RNAi Recombinant Eukaryotic Expression Vectors of Smyd1 and Establishment of Its Stably Transfected H9c2 Cell Lines

作　　者：李帆[1] 廖四芳[1] 刘华友[1] 鲁建鑫[1] 刘宪楚[1] 刘明[1] 吴秀山[1] 李永青[1] 

机构地区：[1]湖南师范大学蛋白质化学及鱼类发育生物学教育部重点实验室心脏发育研究中心,湖南长沙410081

出　　处：《激光生物学报》2011年第5期657-661,共5页Acta Laser Biology Sinica

基　　金：国家然科学基金项目(30930054;30871417;30971663);国家重点基础研究发展计划项目(2005CB522505);湖南省自然科学基金项目(09JJ5014)

摘　　要：运用RNA干扰技术(RNA interference RNAi)构建pSUPER.retro-Smyd1真核表达质粒,经鉴定后用脂质体法转染H9c2细胞,通过G418筛选出稳定表达pSUPER.retro-Smyd1的细胞系,最后经Western blot及RT-PCR实验鉴定其干扰效果。经鉴定,通过H9c2细胞系构建的pSUPER.retro-Smyd1干扰细胞系的干扰效果显著。因此,本实验成功构建了Smyd1干扰真核表达质粒及其稳定转染的H9c2细胞系,为进一步研究Smyd1基因在心脏发育中的作用奠定了良好的实验基础。The eukaryotic expression vector of pSUPER-Smydl was constructed for RNA interference. After verification, we transfected the plasmids into H9c2 cells by Lipofectamine 2000. After screening by G418, stable transfected cell lines were established to express pSUPER-Smydl. Finally, the interferential effect was demonstrated by Western blot and RT-PCR. In a word, RNAi recombinant eukaryotic expression vector of Smydl was successfully Constructed and the establishment of its stably transfected H9c2 cell lines laid a solid experimental foundation for the further studies on the function of Smydl gene of the heart development.

关 键 词：Smyd1基因 RNAI技术 SIRNA H9c2细胞系 稳定转染 

分 类 号：Q78[生物学—分子生物学]