| SELEX筛选LPS寡核苷酸适配子方法的建立 |
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| 引用本文： | 杨清武,文爱清,吕凤林,陈彩宇.SELEX筛选LPS寡核苷酸适配子方法的建立[J].重庆医学,2008,37(16):1773-1775. |
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| 作者姓名： | 杨清武 文爱清 吕凤林 陈彩宇 |
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| 作者单位： | 第三军医大学大坪医院野战外科研究所神经内科,重庆,400042;第三军医大学大坪医院野战外科研究所神经内科,重庆,400042;第三军医大学大坪医院野战外科研究所神经内科,重庆,400042;第三军医大学大坪医院野战外科研究所神经内科,重庆,400042 |
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| 摘 要： | 目的 建立SELEX筛选LPS的适配子方法,为后续大规模筛选奠定基础.方法 在成功构建含40个随机序列的单链DNA(ssDNA)文库的基础上,优化了扩增为双链DNA(dsDNA)文库的PCR反应条件;同时对液相筛选与固液相筛选两种方法进行比较,确定最优的筛选方法并进行4轮筛选.结果 确定了扩增为双链DNA(dsDNA)文库的PCR反应的最佳条件,采用液相筛选方法经过4轮筛选,随机单链DNA(ssDNA)文库与内毒素(LPS)的结合率明显上升,CPM值与初始库相比差异有统计学意义.结论 建立了SELEX筛选LPS寡核苷酸适配子的筛选方法.
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| 关 键 词： | 随机ssDNA文库 指数富集配基的系统进化(SELEX) 寡核苷酸适配子 LPS |
A procedure for SELEX screening aptamers of LPS from ssDNA random library |
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| YANG Qing-wu,WEN Ai-qing,LU Feng-lin,et al..A procedure for SELEX screening aptamers of LPS from ssDNA random library[J].Chongqing Medical Journal,2008,37(16):1773-1775. |
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| Authors: | YANG Qing-wu WEN Ai-qing LU Feng-lin |
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| Abstract: | Objective To construct the method for systematic evolution of ligands by exponential enrichment(SELEX) screening aptamers of LPS from ssDNA random library and to provide the foundation for large scale screening.Methods A 84 nucleotides ssDNA pool containing 40 random nucleotides flanked by invariant primer was designed.The PCR reaction conditions were optimized for converting the ssDNA pool into dsDNA pool.Compared to liquid phase screening and solid phase screening,the optmizing method was determined and 4 rounds selections were done.Results The optmizing PCR amplification for screening condition for converting the ssDNA pool into dsDNA pool was determined.After 4 rounds selections by liquid phase method,the combination percentage of the ssDNA pool bound to LPS inceased.CPM value had signifcant difference compared with initiate pool.Conclusion A procedure for SELEX screening aptamers of LPS from ssDNA random library is constructed. |
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| Keywords: | LPS |
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