| miR-1腺病毒载体的构建及表达分析 |
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| 引用本文： | 许旭东,宋晓伟,李青,林丽,袁文俊,荆清,秦永文.miR-1腺病毒载体的构建及表达分析[J].南昌大学学报(医学版),2010,50(8). |
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| 作者姓名： | 许旭东 宋晓伟 李青 林丽 袁文俊 荆清 秦永文 |
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| 作者单位： | 许旭东,宋晓伟,秦永文,XU Xu-dong,SONG Xiao-wei,QIN Yong-wen(第二军医大学长海医院心血管内科,上海,200433);李青,LI Qing(中国科学院上海生命科学研究院健康科学研究所,上海,200025);林丽,袁文俊,LIN Li,YUAN Wen-jun(第二军医大学生理学教研室,上海,200433);荆清,JIN Qing(第二军医大学长海医院心血管内科,上海,200433;中国科学院上海生命科学研究院健康科学研究所,上海,200025) |
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| 摘 要： | 目的构建并鉴定microRNA-1(miR-1)的腺病毒表达载体。方法 PCR扩增含大鼠miR-1前体的DNA片断,并将其克隆到腺病毒穿梭质粒pAdTrack。pAdTrack经PmeI酶切线性化后与腺病毒骨架质粒pAdEasy-1在BJ5183菌中进行同源重组。重组质粒pAdprecusor-miR-1线性化后,转染293A细胞,进行病毒包装,得到重组腺病毒颗粒Ad-miR-1。Ad-miR-1与Ad-GFP病毒转染培养的乳鼠心肌细胞,通过实时荧光定量PCR方法检测miR-1的表达效率。结果基因测序及酶切鉴定证实重组Adprecursor-miR-1腺病毒载体构建成功;腺病毒Ad-miR-1转染心肌细胞后,实时荧光定量PCR方法证实腺病毒Ad-miR-1能够显著提高心肌细胞内miR-1的表达水平。结论利用同源重组的方法构建的miR-1腺病毒能够提高心肌细胞内miR-1的表达水平。
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| 关 键 词： | 心肌细胞 腺病毒 |
Establishment and Expression Analysis of miR-1 Adenovirus Vector |
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| XU Xu-dong,SONG Xiao-wei,LI Qing,LIN Li,YUAN Wen-jun,JIN Qing,QIN Yong-wen.Establishment and Expression Analysis of miR-1 Adenovirus Vector[J].Journal of Nanchang University(Medical Science),2010,50(8). |
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| Authors: | XU Xu-dong SONG Xiao-wei LI Qing LIN Li YUAN Wen-jun JIN Qing QIN Yong-wen |
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| Institution: | XU Xu-dong1,SONG Xiao-wei 1,LI Qing2,LIN Li 3,YUAN Wen-jun3,JIN Qing1,2,QIN Yong-wen1(1.Department of Cardiology,Changhai Hospital,the Second Military Medical University,Shanghai 200433,China,2.Institute of Health Sciences,Institutes for Shanghai Biological Sciences,Chinese Academy of Sciences,Shanghai 200025,3.Department of Physiology,China) |
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| Abstract: | Objective To construct and identify heart-enriched microRNAs adenovirus vector containing miR-1 precursor gene.Methods The primers of miR-1 precursor was designed and used for PCR amplification.The products were then cloned into adenovirus shuttle plasmid pAdTrack and linearized by enzyme Pme I;the resultant plasmid was co-transfected into E coli BJ51 83cells with adenovirus backbone plasmid pAdEasy-1 for homologous recombination.Then the recombinant plasmid was identified,1 inearized and packaged with QBI-... |
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| Keywords: | microRNAs miR-1 |
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