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双链DNA(dsDNA)中单碱基凸出结构(bulge structure)具有重要生物学意义,这种结构也是DNA靶向药物的目标部位之一.荧光小分子2-氨基-5,6,7-三甲基-1,8-萘啶(ATMND)能够通过氢键识别胞嘧啶(cytosine),因而对dsDNA中凸出的胞嘧啶表现出明显的特异性结合.与其余三种凸出的碱基相比,ATMND与凸出部位胞嘧啶的结合伴随着ATMND荧光的明显猝灭,因而可以用于胞嘧啶凸出结构的识别.利用解旋温度测量、荧光、圆二色光谱对ATMND和存在胞嘧啶凸出结构的dsDNA相互作用进行了研究.荧光滴定结果表明ATMND和dsDNA中凸出部位未配对的胞嘧啶的结合常数K11=4.8×105M?1.通过对含胞嘧啶凸出结构的dsDNA与ATMND结合前后的解旋温度曲线进行解析,发现胞嘧啶凸出结构相邻碱基对凸出的胞嘧啶与ATMND的结合有较大的影响.荧光测量结果也表明ATMND荧光的猝灭效率与凸出结构相邻碱基的类型有关,当相邻碱基为鸟嘌呤(guanine,G)时,荧光猝灭效率最高.基于dsDNA中凸出的碱基对ATMND荧光猝灭效率存在明显差异这一现象,设计了探针DNA实现了乳腺癌相关基因(PGR gene rs3740753)中单核苷酸多态性(G/C变异)的荧光分型.
The single-base bulge structure of double-stranded DNA (dsDNA) is of important biological significance and is also one of the target sites for DNA targeting drugs. The fluorescent small molecule 2-amino-5,6,7- Trimethyl-1,8-naphthyridine (ATMND) was able to recognize cytosine by hydrogen bonding and thus showed a distinct specific binding to the protruding cytosines in dsDNA. In contrast to the other three protruding bases Compared with the cytosine binding of ATMND with ATMND fluorescence quenching, ATMND can be used to recognize the cytosine protuberance.Using spin-on temperature measurement, fluorescence and circular dichroism spectroscopy, Pyrimidine protruding structure dsDNA interactions were studied.Fluorescent titration results showed that ATMND and dsDNA uncoordinated cytosine binding constant K11 = 4.8 × 105M? 1. By containing cytosine protruding structure of dsDNA and ATMND before and after the unwinding temperature curve analysis and found that the protruding cytosine protruding base adjacent to the protruding cytosine ATMND combination has a greater impact.Fluorescence measurements also showed that ATMND fluorescence quenching efficiency and convex The structure of adjacent bases is related to the type of base when adjacent to the base Is guanine (G), the fluorescence quenching efficiency is highest.According to the obvious difference of the fluorescent quenching efficiency of ATMND in the protruding base of dsDNA, the probe DNA is designed to realize the expression of breast cancer related gene (PGR gene rs3740753) single nucleotide polymorphism (G / C mutation) of the fluorescent typing.