Beijing Advanced Innovation Center for Materials Genome Engineering , University of Science and Technology Beijing , 30 Xueyuan Road , Beijing 100083 , P.R. China.
Beijing Key Laboratory for Bioengineering and Sensing Technology, Research Center for Bioengineering and Sensing Technology , School of Chemistry & Biological Engineering University of Science & Technology Beijing , 30 Xueyuan Road , Beijing 100083 , P.R. China.
Anal Chem. 2019 Aug 6;91(15):9828-9835. doi: 10.1021/acs.analchem.9b01487. Epub 2019 Jul 18.
There is intense interest in visualizing intracellular microRNA (miRNA) in situ due to its valuable diagnostic and prognostic function and expressional heterogeneity. The direct monitoring of intracellular miRNA expressional level with a wide concentration range and in ultralow amount remains a key challenge. Herein, we report a responsive DNA "nano wheel" (DNW) by hybridization of modified six DNA hairpin structures to a programmed DNA nanowire for highly efficient and fast miRNA imaging in living cells. We demonstrate that one target miRNA is able to trigger the nearby DNA hairpins sequential hybridization through accelerated localized DNA cascade reaction (LDCR) along the DNA nanowire and open of the self-quenched hairpins to assemble six-arm branched junction "nano wheel" structure lighted with three fluorescence dye. Notably, the recycled miRNA target further acts as an catalyst to generate multiple lighted "nano wheel" structures, thus one target miRNA enable to trigger strong and amplified fluorescence signal for intracellular sensitive imaging. Meanwhile, the LDCR endows the system with fast reaction kinetics owing to the high local concentration. It outperforms the "one-to-one" conventional molecular beacon (MB) and intermolecular hybridization-based six-arm branched junction in sensitivity and reaction kinetics for intracellular miRNA imaging verified using MCF-7, A549, Hela, and NHDF cells. The programmed DNW provides a useful and valuable tool for rapid and highly efficient visualization of low-abundance miRNA in living cells, which is beneficial to understand the miRNA function and exploration of its biomedical application.
由于其有价值的诊断和预后功能以及表达异质性，人们对在体内可视化细胞内 microRNA（miRNA）产生了浓厚的兴趣。由于其广泛的浓度范围和超低的含量，直接监测细胞内 miRNA 表达水平仍然是一个关键挑战。在此，我们通过杂交修饰的六个 DNA 发夹结构来报告一种响应性 DNA“纳米轮”（DNW），以程序化的 DNA 纳米线，用于在活细胞中进行高效快速的 miRNA 成像。我们证明，一个靶 miRNA 能够通过沿 DNA 纳米线加速局部 DNA 级联反应（LDCR），触发附近 DNA 发夹的顺序杂交，并打开自我淬灭的发夹以组装带有三个荧光染料的六臂分支结“纳米轮”结构。值得注意的是，循环 miRNA 靶标进一步充当催化剂，以产生多个点亮的“纳米轮”结构，从而一个靶标 miRNA 能够触发强烈和放大的荧光信号，用于细胞内敏感成像。同时，LDCR 由于高局部浓度赋予了系统快速的反应动力学。它在 MCF-7、A549、Hela 和 NHDF 细胞中用于细胞内 miRNA 成像的灵敏度和反应动力学方面，优于“一对一”传统分子信标（MB）和基于分子间杂交的六臂分支结。程序化的 DNW 为快速、高效地可视化活细胞中低丰度 miRNA 提供了一种有用且有价值的工具，这有利于了解 miRNA 的功能及其在生物医学中的应用探索。
