生命科学研究2011，Vol.15Issue(2)：119-124,6.
靶向CrkL基因的siRNA真核表达载体的构建及对H9C2心肌细胞凋亡的作用
Construction of Eukaryotic siRNA Expression Vector Targeting CrkL gene and Its Effect on Appoptosis in H9C2 Cells of Rat Cardiomycytes
摘要
Abstract
To provide an experimental basis for studying CrkL gene action mechanism in Cardiomyocyte.First, six antisense oligodeoxynucleotide sequences targeting rat's CrkL mRNA were designed and synthesized in vitro, and then directionally cloned into pSR-GFP/neo siRNA expressing plasmid respectively.The recombinant vectors pSR-GFP/neo-CrkLi were constructed and identified successfully by double restriction endonuclease digestion analysis and DNA sequencing.The pSR-GFP/neo-CrkLi were transfected into H9C2 cell line of rat cardiomyocyte, and the transfection efficiency was evaluated with fluorescence microscope.The inhibitive effects of the recombinants on CrkL mRNA were observed by RT-PCR.And the effects of CrkL gene on appoptosis of H9C2 cells was detected by flow cytometry.Six pSR-GFP/neo-CrkLi were transfected into H9C2 cell efficiently.The expression of CrkL mRNA in H9C2 cells transfected with pSRGFP/neo-CrkLi was significantly inhibited in three vectors.Flow cytometry showed that the apoptosis rate of group transfected with pSR-GFP/neo-CrkLi-3 was significantly higher than the other groups(P=0.001 ).关键词
CrkL基因/RNA干扰/心肌细胞/H9C2细胞株Key words
CrkL/ RNA interference/ cardiomyocyte/ H9C2 cell line分类
生物科学引用本文复制引用
朱炳豹,李刚..靶向CrkL基因的siRNA真核表达载体的构建及对H9C2心肌细胞凋亡的作用[J].生命科学研究,2011,15(2):119-124,6.基金项目
重庆市卫生局医学科学技术研究项目(2009-2-290,04-2-154) （2009-2-290,04-2-154）
重庆市科委自然科学基金计划资助项目(CSTC,2007BB5276) （CSTC,2007BB5276）
