The RNA Institute, University at Albany, State University of New York, Albany, NY 12222, USA.
Department of Biological Sciences, University at Albany, State University of New York, Albany, NY 12222, USA.
Sci Adv. 2019 Mar 13;5(3):eaau9443. doi: 10.1126/sciadv.aau9443. eCollection 2019 Mar.
MicroRNAs are short noncoding regulatory RNAs that are increasingly used as disease biomarkers. Detection of microRNAs can be arduous and expensive and often requires amplification, labeling, or radioactive probes. Here, we report a single-step, nonenzymatic microRNA detection assay using conformationally responsive DNA nanoswitches. Termed miRacles (microRNA-activated conditional looping of engineered switches), our assay has subattomole sensitivity and single-nucleotide specificity using an agarose gel electrophoresis readout. We detect cellular microRNAs from nanogram-scale RNA extracts of differentiating muscle cells and multiplex our detection for several microRNAs from one biological sample. We demonstrate 1-hour detection without expensive equipment or reagents, making this assay a compelling alternative to quantitative polymerase chain reaction and Northern blotting.
微 RNA 是短的非编码调控 RNA，越来越多地被用作疾病生物标志物。微 RNA 的检测可能很困难和昂贵，并且通常需要扩增、标记或放射性探针。在这里，我们报告了一种使用构象响应性 DNA 纳米开关的单步非酶微 RNA 检测方法。我们的方法称为 miRacles（微 RNA 激活的工程开关的条件环化），使用琼脂糖凝胶电泳读出，具有亚皮摩尔灵敏度和单核苷酸特异性。我们从分化的肌肉细胞的纳克级 RNA 提取物中检测细胞微 RNA，并从一个生物样本中对多个微 RNA 进行多重检测。我们证明了无需昂贵的设备或试剂即可在 1 小时内进行检测，这使得该检测方法成为定量聚合酶链反应和 Northern 印迹的有力替代品。
