| Cyr61基因siRNA表达载体的构建及沉默作用研究 |
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| 引用本文： | 张燕,武庆平,姚尚龙.Cyr61基因siRNA表达载体的构建及沉默作用研究[J].细胞与分子免疫学杂志,2011,27(4):360-363. |
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| 作者姓名： | 张燕 武庆平 姚尚龙 |
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| 作者单位： | 华中科技大学同济医学院附属协和医院麻醉科,湖北,武汉,430022 |
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| 基金项目： | 国家自然科学基金资助项目(30930089) |
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| 摘 要： | 目的:构建Cyr61靶向siRNA重组表达载体,为探讨抑制Cyr61基因表达对机械通气所致肺损伤的研究奠定基础。方法:根据GenBank数据库提供的Cyr61基因核苷酸序列,选择设计3条能转录siRNA的DNA序列,命名Cyr61-1 siRNA,Cyr61-2 siRNA,Cyr61-3 siRNA,同时设计1条非特异性序列作为阴性对照。据此设计合成各自的寡核苷酸链,退火后连接入pGenesil1.1载体,转化扩增后进行序列测定。4种重组表达载体转染肺癌A549细胞,逆转录RT-PCR和Western blot法分别在mRNA和蛋白水平检测Cyr61的表达。结果:经酶切鉴定和测序结果证实Cyr61靶向siRNA重组表达载体构建成功,它对大肠癌细胞Cyr61 mRNA和蛋白的表达抑制率分别为60.54%和52.97%。结论:Cyr61靶向siRNA重组表达载体构建成功并能显著抑制Cyr61基因的表达。
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| 关 键 词： | Cyr61 RNA干扰 基因治疗 |
Construction of Cyr61 siRNA expression vector and its Characteristic investigation |
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| ZHANG Yan,WU Qing-ping,YAO Shang-long.Construction of Cyr61 siRNA expression vector and its Characteristic investigation[J].Journal of Cellular and Molecular Immunology,2011,27(4):360-363. |
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| Authors: | ZHANG Yan WU Qing-ping YAO Shang-long |
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| Institution: | ZHANG Yan,WU Qing-ping,YAO Shang-long Department of Anesthesiology,Union Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430022,China |
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| Abstract: | AIM:To construct si RNA recombinant expression vector targeting Cyr61 gene,and to screen the stably transfected cell clone.METHODS: Three si RNAs sequences based on the sequence of Cyr61mRNA in the GenBank were designed and synthesized,and one scrambled siRNA sequences served as negative control.The cDNA was synthesized and inserted into plasmid pGenesil1.1.The plasmids was sequenced to confirm the inserted sequence.RT-PCR and Western blot analysis was used to assess the levels of Cyr61 mRNA and proteins af... |
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| Keywords: | Cyr61 RNA interference gene therapy |
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