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| 串联MS2衣壳蛋白和MS2特异结合RNA表达载体构建及应用 | |
| 引用本文： | 蒋太峰,付汉江,刘珊珊,金英花,朱捷,李恩民,郑晓飞.串联MS2衣壳蛋白和MS2特异结合RNA表达载体构建及应用[J].军事医学科学院院刊,2012,36(8):595-598. |
| 作者姓名： | 蒋太峰 付汉江 刘珊珊 金英花 朱捷 李恩民 郑晓飞 |
| 作者单位： | 1. 军事医学科学院放射与辐射医学研究所,北京,100850;汕头大学医学院,汕头,515041 2. 军事医学科学院放射与辐射医学研究所,北京,100850 3. 军事医学科学院放射与辐射医学研究所,北京,100850;吉林大学生命科学学院,长春,130012 4. 吉林大学生命科学学院,长春,130012 5. 汕头大学医学院,汕头,515041 |
| 基金项目： | 国家重大科学研究计划资助项目，国家重点基础研究发展计划(973计划)资助项目，国家自然科学基金资助项目，北京市自然科学基金资助项目 |
| 摘 要： | 目的构建体内研究RNA-蛋白相互作用的表达载体。方法利用MS衣壳蛋白可与MS2结合位点（MS2bs）RNA特异性结合的特点，通过设计特异引物，构建pcDNA3．0-Flag．2XMS2和pcDNA3．0—12XMS2％s表达载体，以及表达MEG3非编码RNA的表达载体pcDNA3．0-EG3V1—12XMS2bs。共转染细胞，进行RNA免疫沉淀，所得RNA进行实时定量PCR分析验证。结果成功构建了pcDNA3．0-Flag-2XMS2和pcDNA3．0．12XMS2bs表达载体，实时定量PCR结果表明，与对照相比能明显富集MEG3V1非编码RNA分子。结论成功构建了体内研究RNA-蛋白质相互作用的表达载体，为RNA一蛋白相互作用研究提供了新的技术方法。 |
| 关 键 词： | MS2衣壳蛋白 RNA-蛋白相互作用 RNA结合蛋白 |
Construction and application of multi-MS2 coat protein expression vector and multi MS2 specific binding RNA expression vector |
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| JIANG Tai-feng , FU Han-jiang , LIU Shan-shan , JIN Ying-hua , ZHU Jie , LI En-min , ZHENG Xiao-fei.Construction and application of multi-MS2 coat protein expression vector and multi MS2 specific binding RNA expression vector[J].Bulletin of the Academy of Military Medical Sciences,2012,36(8):595-598. | |
| Authors: | JIANG Tai-feng FU Han-jiang LIU Shan-shan JIN Ying-hua ZHU Jie LI En-min ZHENG Xiao-fei |
| Institution: | ( I. Institute of Radiation Medicine, Academy of Military Medical Sciences, Beijing 100850, China; 2. Medical College, Shantou University ,Shantou 515041 , China; 3. College of Life Sciences, ]ilia University, Changchun 130012, China) Co-corresponding authors, ZHENG Xiao-fei, Tel ：010-66931237, E-mail ： zhengxf@ nic. bmi. ac. cn ; LI En-min, Tel ：0754-88900413, E-mail： nmli@ stu. edu. cn |
| Abstract: | Objective To construct two kinds of novel expression vectors for RNA-protein interaction studies in vivo. Methods The pcDNA3.0-Flag-2XMS2 vector containing two repeat MS2 coat protein sequence and the pcDNA3.0- 12XMS2bs vector containing 12 repeat MS2bs（MS2 binding sites） RNA sequence were constructed. The pcDNA3. O- MEG3VI-12XMS2bs vector expressing non-coding RNA MEG3 V1 was also constructed, pcDNA3.0-Flag-2XMS2 vector and pcDNA3.0-MEG3VI-12XMS2bs vector were co-transfected into HEK293 cells. The level of MEG3 V1 RNA from co- transfected cells was detected by RNA immunoprecipitation and real-time PCR. Results pcDNA3.0-Flag-2XMS2 vector and pcDNA3.0-12XMS2bs vector were constructed successfully. Real-time PCR results showed that RNA immunoprecipitati- on could significantly enrich MEG3V1 RNA. Conclusion We have successfully constructed expression vectors for RNA-pro- rein interaction studies in vivo. |
| Keywords: | MS2 coat protein RNA-protein interaction RNA-binding proteins |
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