基于金纳米粒子的催化发夹DNA组装检测microRNA

摘要

By combining gold nanoparticles (AuNPs) with catalytic hairpin DNA assembly,we develop a new method for the detection of microRNA (miRNA).Firstly,fluorescein-labeled hairpin DNA probe P1 was modified on the surface of AuNPs.The fluorescence signal of P1 is quenched by the AuNPs.When the target miRNA is present,it can hybrid with P1 and form the P1-miRNA intermediates.At the same time,the hairpin structure of P1 is opened.Then the opened P1 can hybrid with the hairpin P2 to form a stable structure,P1-P2,while the fluorophore in P1 is far away from the AuNPs,resulting in the fluorescence emission of fluorescein.Meanwhile,the target miRNA is released and can subsequently induce the multiple assembled reactions of P1 and P2 on the surface of AuNPs.By repeating the cycle,the amplified detection of miRNA is achieved.The method shows a good linear relationship with the miRNA in the concentration range of 50 pmol/L to 2 nmol/L.The detection limit was estimated to be 39 pmol/L.The method is simple,low cost,and used for the detection of miRNA in HepG 2 cells and HeLa cells with satisfactory results.%结合金纳米粒子和催化发夹DNA组装,发展了一种检测microRNA(miRNA)的新方法.首先,在金纳米粒子表面修饰荧光素标记的发夹DNA探针P1,P1荧光被金纳米粒子猝灭.当加入目标miRNA分子时,其与P1杂交并形成P1-miRNA中间体,同时打开P1发夹结构.继而P1-miRNA与P2发夹探针结合,形成稳定的P1-P2双链结构,导致P1上的荧光素远离金纳米粒子而释放荧光,同时顶替下miRNA.随后,miRNA又可以引发金纳米粒子表面多个P1与P2探针的组装,如此循环,实现了对miRNA的放大检测.方法中miRNA在50 pmol/L～2 nmol/L之间呈现良好的线性关系,最低检出限为39 pmol/L.方法简便,成本低,应用于HeLa细胞和HepG 2细胞中miRNA的检测,结果满意.