pcDNA3.0-RKIP重组质粒构建及其在PC12细胞中的表达  

Construction of the recombinant plasmid of pcDNA3.0-RKIP and expression in PC12 cells

作　　者：林涛[1] 左红艳[1] 王德文[1] 彭瑞云[1] 周培岚[2] 王水明[1] 高亚兵[1] 王少霞[1] 

机构地区：[1]军事医学科学院放射与辐射医学研究所实验病理学研究室,北京100850 [2]军事医学科学院毒物药物研究所,北京100850

出　　处：《细胞与分子免疫学杂志》2011年第4期399-401,404,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目(30770527);国家科技重大专项(2008ZXJ09004-019)

摘　　要：目的:构建RKIP真核表达质粒,并检测其在PC12细胞中的表达。方法:从大鼠背根神经节细胞中提取总RNA,应用巢式PCR技术,扩增获得RK IP基因编码序列片段,克隆入真核表达载体pcDNA3.0,对重组质粒进行酶切和测序鉴定后,以Vigofect非脂质体介导法转染至PC12细胞,通过Western blot检测RKIP蛋白的表达。结果:酶切和测序结果证明pcDNA3.0-RKIP重组质粒的DNA序列完全正确,将其转染PC12细胞后,RKIP蛋白表达明显增加。结论:pcDNA3.0-RKIP真核表达质粒构建成功,并能在PC12细胞内表达,为深入研究RKIP的神经生物学功能提供了实验基础。AIM：To construct the eukaryotic expression vectors of RKIP plasmid and detect its expression in PC12 cells.METHODS： The coding sequence of RKIP was generated by nested-PCR using total RNA extracted from the root ganglion neurons of rats.RKIP gene was cloned into the eukaryotic expression vector pcDNA3.0.After restriction enzyme analysis and sequence identification,the recombinant plasmid was transfected into PC12 cells with non-liposome mediated method by Vigofect.The expression of RKIP was detected by Western blot.RESULTS： The results of enzyme analysis and sequencing both identified DNA sequence of recombinant plasmid pcDNA3.0-RKIP correctly.The expression of RKIP increased obviously after transfection into PC12 cells.CONCLUSION： The eukaryotic expression plasmid of pcDNA3.0-RKIP was constructed successfully and it can be sustainly expressed in PC12 cells.This provides experimental basis for further study on the neurological function of RKIP.

关 键 词：RKIP 重组质粒 基因转染 PC12细胞 

分 类 号：Q684[生物学—生物物理学]