一种高效灵敏的DNA荧光探针的合成及应用

Synthesis and Application of an Efficient and Sensitive DNA Fluorescent Probe

设计合成了一种新型噻唑橙二聚体荧光染料Bi-TO3,采用荧光发射光谱、圆二色光谱及活细胞荧光成像等方法研究了其与DNA的相互作用.在10 mmol/L Tris-HCl缓冲液(pH=7.4)中,Bi-TO3的固有荧光极弱,量子产率小于0.001%;与小牛胸腺DNA结合后,其荧光可显著增强约950倍,但对RNA和蛋白等生物大分子及黏度等环境因素则无明显响应.紫外吸收光谱及圆二色光谱滴定实验表明,Bi-TO3以小沟结合模式与DNA作用,且对AT序列有选择性.实验结果表明,在缓冲溶液中Bi-TO3的荧光增强信号与低浓度范围的poly(dA-dT)2仍呈良好的线性关系,检出限为13.3 ng/mL,灵敏较度高;且Bi-TO3可在较低浓度范围(6～12μmol/L)内应用于活细胞荧光成像.

A novel thiazole orange dimmer derivative Bi-TO3 was developed.The interaction of Bi-TO3 with DNA was investigated through the absorption,emission,circular dichroism spectroscopy and live-cell imaging experiments.As a specific probe for DNA,Bi-TO3 itself was almost non-fluorescent（quantum yield〈0.001%） in buffer.Upon binding to DNA,the fluorescence of Bi-TO3 increased approximately 950-fold,even without interferences of responding to RNA,BSA,viscosity,etc.Absorption and circular dichroism spectra showed that Bi-TO3 associated with DNA in a minor groove binding mode.The emission detection limit was 13.3 ng/mL,which manifested that Bi-TO3 could detect trace DNA in solution.Moreover,Bi-TO3 could be applied for live-cell fluorescence imaging with relatively low concentrations of 6—12 μmol/L.