Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, School of Chemistry and Materials Science, Shaanxi Normal University, Xi'an 710062, China.
Appl Spectrosc. 2012 Feb;66(2):170-4. doi: 10.1366/11-06410.
The fluorescent small molecule 2-amino-7-methyl-1,8-naphthyridine (AMND) can selectively bind to a cytosine (C) at a C-C mismatch in double-stranded DNA (dsDNA). The interactions between AMND and C-C mismatch-containing dsDNA were investigated by measuring ultraviolet (UV) absorption as a function of temperature to obtain melting curves as well as circular dichroism and fluorescence spectra. Results show that AMND strongly stabilizes C-C mismatch-containing dsDNA, whereas fully matched duplexes are not stabilized under the same conditions. The fluorescence of AMND was efficiently quenched when it was bound to a C-C mismatch in dsDNA. Binding constants (K(11)), obtained by fluorescence titration, were 1.2 × 10(5) M(-1). Although sensing functions depend on the sequences flanking the mismatch site, the change in AMND fluorescence intensity can be utilized to detect the C-C mismatch-containing dsDNA. Accordingly, discrimination of the C/G mutation in the model sequence (PGR gene rs1255998) was achieved by visualizing fluorescence of AMND. A probe DNA molecule was designed to contain a C opposite the C/G base in the target DNA, and this probe was used to hybridize the target DNA. The fluorescence of AMND was "on" for a C-G match, while the fluorescence was "off" for a C-C mismatch. This assay is simple and does not require DNA labeling.
荧光小分子 2-氨基-7-甲基-1,8-萘啶（AMND）可以选择性地与双链 DNA（dsDNA）中的胞嘧啶（C）在 C-C 错配处结合。通过测量紫外（UV）吸收随温度的变化来研究 AMND 与含 C-C 错配的 dsDNA 之间的相互作用，以获得熔解曲线以及圆二色性和荧光光谱。结果表明，AMND 强烈稳定含 C-C 错配的 dsDNA，而在相同条件下，完全匹配的双链体则不会被稳定。当 AMND 与 dsDNA 中的 C-C 错配结合时，其荧光被有效猝灭。通过荧光滴定获得的结合常数（K（11））为 1.2×10(5) M(-1)。尽管传感功能取决于错配位点侧翼的序列，但 AMND 荧光强度的变化可用于检测含 C-C 错配的 dsDNA。因此，可以通过可视化 AMND 的荧光来实现对模型序列（PGR 基因 rs1255998）中 C/G 突变的区分。设计了一个探针 DNA 分子，使其在靶 DNA 中与 C 相对应的位置含有 C，并用该探针杂交靶 DNA。AMND 的荧光对于 C-G 匹配为“开”，而对于 C-C 错配为“关”。该测定方法简单，不需要 DNA 标记。
