Ultrafast capillary electrophoresis isolation of DNA aptamer for the PCR amplification-based small analyte sensing.
Ultrafast capillary electrophoresis isolation of DNA aptamer for the PCR amplification-based small analyte sensing.
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DOI：
10.3389/fchem.2015.00049
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发表时间：
2015
影响因子：
5.5
通讯作者：
Ravelet C
中科院分区：
文献类型：
--
作者：
Fiore E;Dausse E;Dubouchaud H;Peyrin E;Ravelet C
在这里，我们报告了一种新的基于均质 DNA 扩增的适体检测，用于小分析物传感。选择作为模型分析物的腺苷适体被分成两个片段，能够在靶标存在的情况下组装。在一个片段的末端引入引物，以产生可扩增的 DNA 成分。可扩增片段的量可通过实时聚合酶链式反应 (RT-PCR) 扩增进行定量，并直接可靠地依赖于腺苷浓度。该方法结合了毛细管电泳 (CE) 的极高分离效率和同质形式（无需固定）以及实时 PCR 扩增的灵敏度。通过设计 CE 输入/输出方案，开发了超快速分离目标结合的分裂适体 (60 s)。该方法成功应用于LOD为1 μM的腺苷的测定。
Here, we report a new homogeneous DNA amplification-based aptamer assay for small analyte sensing. The aptamer of adenosine chosen as the model analyte was split into two fragments able to assemble in the presence of target. Primers were introduced at extremities of one fragment in order to generate the amplifiable DNA component. The amount of amplifiable fragment was quantifiable by Real-Time Polymerase Chain Reaction (RT-PCR) amplification and directly reliable on adenosine concentration. This approach combines the very high separation efficiency and the homogeneous format (without immobilization) of capillary electrophoresis (CE) and the sensitivity of real time PCR amplification. An ultrafast isolation of target-bound split aptamer (60 s) was developed by designing a CE input/ouput scheme. Such method was successfully applied to the determination of adenosine with a LOD of 1 μM.