基于荧光标记核酸探针及核酸染料SYBR GreenⅠ定量检测单链DNA

Quantitative Determination of Sequence-specific Nucleic Acid Based on Fluorescent Labeled DNA Probe and Dye SYBR Green Ⅰ

建立了同步双色荧光定量检测单链DNA的方法。利用氧化石墨烯将荧光标记核酸探针的荧光猝灭,而当其与待测液中的目标DNA发生杂交反应,生成的双链DNA脱离氧化石墨烯而使得染料荧光得以恢复,同时生成的双链DNA与核酸染料SYBR GreenⅠ结合,使荧光增强。在优化条件下,目标DNA浓度在1×10"10～2×10"9mol/L范围内时,两种荧光染料的荧光强度之和与目标DNA的浓度之间具有良好的线性关系,拟合的回归方程为y=63.388x+9.3862,R2=0.9954,检出限为85 pmol/L。本方法灵敏度高、准确性及选择性好。

A synchronous fluorescence spectral method was developed for the quantitative detection of sequence-specific DNA. Graphene oxide was used to quench the fluorescence of the dyes. When the target DNA was added, the fluorescences of the two dyes were recovered due to the formed DNA duplex was far away from graphene oxide. Under the optimum conditions, the linear range was 1× 10^-10 -2 × 10^-9 mol/L, the calibration curve was y = 63. 388x＋9. 3862, R^2 = 0. 9954. The detection limit of 85 pmol/L was estimated with 3σ （where σ was the relative standard deviation of a blank solution, n = 11 ）. This method has good sensitivity, selectivity and accuracy.