Use of vitamin B2 for fluorescence detection of thymidine-related single-nucleotide polymorphisms |
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Authors: | Nishizawa Seiichi Sankaran N B Seino Takehiro Cui Ying-Yu Dai Qing Xu Chun-Yan Yoshimoto Keitaro Teramae Norio |
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Institution: | a Department of Chemistry, Graduate School of Science, Tohoku University, Aoba-ku, Sendai 980-8578, Japan b CREST, Japan Science and Technology Agency (JST), Sendai 980-8578, Japan |
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Abstract: | In combination with abasic site (AP site)-containing DNAs, potential use of a biotic fluorescence compound, Vitamin B2 (riboflavin), is demonstrated for the fluorescence detection of the thymine (T)-related single-nucleotide polymorphisms. Our method is based on construction of the AP site in DNA duplexes, which allows small ligands to bind to target nucleotides accompanied by fluorescence signaling: an AP site-containing probe DNA is hybridized with a target DNA so as to place the AP site toward a target nucleobase, by which hydrophobic microenvironments are provided for ligands to recognize target nucleotides through stacking and hydrogen-bonding interactions. In 10 mM sodium cacodylate buffer solutions (pH 7.0) containing 100 mM NaCl and 1.0 mM EDTA, Vitamin B2 is found to selectively bind to T (K11 = 1.8 × 106 M−1 at 5 °C) over other nucleobases, and this is accompanied by significant quenching of its fluorescence. While the sensing functions depend on the flanking sequences to the AP site, Vitamin B2 is applicable to the detection of T/C (cytosine), T/G (guanine) and T/A (adenine) mutation sequences of the CYP2A6 gene, where the flanking nucleobases are guanines in both positions (-GXG-, X = AP site). |
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Keywords: | Vitamin B2 Abasic site Ligand Hydrogen bond Stacking Fluorescence detection Single-nucleotide polymorphism |
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