首页 | 本学科首页   官方微博 | 高级检索  
     

小鼠微RNA miR-21真核表达质粒的构建及其在小鼠肾小球系膜细胞中的表达
引用本文:张政,彭惠民,徐晓明,何晓燕.小鼠微RNA miR-21真核表达质粒的构建及其在小鼠肾小球系膜细胞中的表达[J].粉末涂料与涂装,2010,23(7).
作者姓名:张政  彭惠民  徐晓明  何晓燕
作者单位:张政,ZHANG Zheng(重庆医科大学基础医学院细胞生物学及遗传学教研室,重庆,400016;重庆医科大学分子医学及肿瘤研究中心,重庆,400016);彭惠民,徐晓明,何晓燕,PENG Hui-min,XU Xiao-ming,HE Xiao-yan(重庆医科大学基础医学院细胞生物学及遗传学教研室,重庆,400016) 
摘    要:目的构建小鼠微RNA miR-21真核表达质粒,并在小鼠肾小球系膜细胞中表达。方法人工合成小鼠miR-21基因序列,构建miR-21真核表达质粒pGenesil-miR-21。使用脂质体Lipofectamine2000转染小鼠肾小球系膜细胞后,G418筛选,获得稳定转染克隆,提取总RNA,通过实时荧光定量RT-PCR技术检测miR-21的表达。结果经酶切鉴定和测序证实,合成的miR-21基因序列完全正确,并已成功克隆到真核表达质粒pGenesil-1上。重组真核表达质粒转染小鼠肾小球系膜细胞后,筛选出的阳性克隆可稳定高表达miR-21。结论已成功构建miR-21真核表达质粒,并在小鼠肾小球系膜细胞中高效表达,为进一步探讨miR-21的生物学功能奠定了基础。

关 键 词:微RNA  miR-21  真核细胞  基因表达  肾小球系膜细胞

Construction of Eukaryotic Expression Vector for Mouse MicroRNA miR-21 and Its Expression in Mouse Mesangial Cells
ZHANG Zheng,PENG Hui-min,XU Xiao-ming,HE Xiao-yan.Construction of Eukaryotic Expression Vector for Mouse MicroRNA miR-21 and Its Expression in Mouse Mesangial Cells[J].Chinese Journal of Biologicals,2010,23(7).
Authors:ZHANG Zheng  PENG Hui-min  XU Xiao-ming  HE Xiao-yan
Abstract:Objective To construct the eukaryotic expression vector for mouse microRNA miR-21 and express in mouse mesangial cells.Methods The miR-21 gene was chemically synthesized and cloned into expression vector pGenesil-1.The constructed recombinant plasmid pGenesil-miR-21 was transfected to mouse mesangial cells in mediation of Lipofectamine 2000,and the stably transfected clones were screened with G418,from which total RNA was extracted for determination of expression of miR-21 by real-time fluorescent quantitative RT-PCR technique.Results Both restriction analysis and sequencing proved that recombinant plasmid pGenesil-miR-21 was constructed correctly.The miR-21 gene was stably and highly expressed in the screened positive clones of mouse mesangial cells transfected with the constructed recombinant plasmid.Conclusion The eukaryotic expression vector for miR-21 was successfully constructed and highly expressed in mouse mesangial cells,which laid a foundation of further investigation of biological function of miR-21.
Keywords:miR-21
本文献已被 万方数据 等数据库收录!
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号