Fluorescence detection of cytosine/guanine transversion based on a hydrogen bond forming ligand
Fluorescence detection of cytosine/guanine transversion based on a hydrogen bond forming ligand
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DOI：
10.1016/j.talanta.2003.09.027
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发表时间：
2004-05-10
期刊：
影响因子：
6.1
通讯作者：
Teramae, N
中科院分区：
文献类型：
--
作者：
Nishizawa, S;Yoshimoto, K;Teramae, N
与含脱碱基位点（AP位点）的寡脱氧核苷酸（ODN）相结合，我们证明了氢键形成配体2-氨基-7-甲基-1,8-萘啶（AMND）的潜在用途，可用于癌症抑制基因p53的胞嘧啶（C）/鸟嘌呤（G）突变序列的荧光检测。我们的方法基于ODN双链体中AP位点的构建，它允许小的合成配体与目标核碱基结合并伴随荧光信号传导：含有AP位点的ODN与目标ODN杂交，从而将AP位点朝向目标核碱基，从而为配体提供疏水微环境以通过氢键识别目标核碱基。在含有 100 mM NaCl 和 1.0 mM EDTA 的 10 mM 二甲胂酸钠缓冲溶液（pH，7.0）中，AMND 与目标 ODN 中的 C（K-d = 1.5 x 10(-6) M）强烈结合，而对 G 的结合亲和力相对适中（K-d = 50 x 10(-6) M）。 AMND仅在与C结合时才会观察到显着的荧光猝灭，使得肉眼判断C/G颠换成为可能。 (C) 2003 Elsevier B.V. 保留所有权利。
in combination with abasic site (AP site)-containing oligodeoxynucleotides (ODNs), we demonstrate potential use of a hydrogen bond forming ligand, 2-amino-7-methyl-1,8-naphthyridine (AMND), for the fluorescence detection of the cytosine (C)/guanine (G) mutation sequence of the cancer repression gene p53. Our method is based on construction of the AP site in ODN duplexes, which allows small synthetic ligands to bind to target nucleobases accompanied by fluorescence signaling: an AP site-containing ODN is hybridized with a target ODN so as to place the AP site toward a target nucleobase, by which hydrophobic microenvironments are provided for ligands to recognize target nucleobases through hydrogen-bonding. In 10 mM sodium cacodylate buffer solutions (pH, 7.0) containing 100 mM NaCl and 1.0 mM EDTA, AMND is found to strongly bind to C (K-d = 1.5 x 10(-6) M) in the target ODN while the binding affinity for G is relatively moderate (K-d = 50 x 10(-6) M). Significant fluorescence quenching of AMND is observed only when binding to C, making it possible to judge the C/G transversion with the naked eye. (C) 2003 Elsevier B.V. All rights reserved.