基于免标记型的DNA甲基化电化学研究
首发时间:2014-11-12
摘要:通过巯基丙酸将DNA探针分子交联在金电极表面,构建了DNA生物传感器。目标DNA与电极表面DNA探针杂交后,利用限制性内切酶HpaII对5'-CCGG-3'序列特异性识别时的甲基化敏感性特点,HpaII会切割未甲基化的dsDNA,而甲基化的dsDNA会禁阻酶切过程。以有机染料亚甲基蓝(MB)为指示剂,通过酶切前后双链DNA链长度变化引起的电流变化,对DNA特异位点的甲基化情况进行定量分析。结果表明,在5×10-6 ~1×10-8 mol L-1浓度范围内,MB在修饰电极上的峰电流的差值与目标DNA链浓度的负对数值呈现出较好的线性关系,线性相关系数为0.9860,回收率为91.16~102.1%。
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Label-free electrochemical research for DNA methylation
Abstract:A label-free DNA biosensor is construced to detect the methylation condition in a specific period of gene based on restrict enzyme. Restrict enzyme HpaII recognizes and digests the specific sites of 5'-CCGG-3'in a dsDNA. Methylene blue (MB) is used as the intercalate indicator to investigate the length variation of the dsDNA before and after enzyme digestion. Restrict enzyme HpaII can cut those dsDNA that are unmethylatted in the specific sites which shorten the length of this dsDNA. And in a result, the quantity of MB that intercalated in the dsDNA decreased and the peak current of MB decreased correspondingly. Meanwhile the methylated DNA remains almost the same after enzyme digestion and the peak currents change little. The relationship is obtained between the degree of a dsDNA and the differential of peak current of MB.
Keywords: electrochemistry DNA methyaltion label-free DNA biosensor
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