Zhang Xinyue, Wang Yong, Fricke Brandon L, Gu Li-Qun
Department of Bioengineering and Dalton Cardiovascular Research Center, University of Missouri , Columbia, Missouri 65211, United States.
ACS Nano. 2014 Apr 22;8(4):3444-50. doi: 10.1021/nn406339n. Epub 2014 Mar 26.
Many efforts are being made in translating the nanopore into an ultrasensitive single-molecule platform for various genetic and epigenetic detections. However, compared with current approaches including PCR, the low throughput limits the nanopore applications in biological research and clinical settings, which usually requires simultaneous detection of multiple biomarkers for accurate disease diagnostics. Herein we report a barcode probe approach for multiple nucleic acid detection in one nanopore. Instead of directly identifying different targets in a nanopore, we designed a series of barcode probes to encode different targets. When the probe is bound with the target, the barcode group polyethylene glycol attached on the probe through click chemistry can specifically modulate nanopore ion flow. The resulting signature serves as a marker for the encoded target. Therefore counting different signatures in a current recording allows simultaneous analysis of multiple targets in one nanopore. The principle of this approach was verified by using a panel of cancer-derived microRNAs as the target, a type of biomarker for cancer detection.
人们正在做出许多努力，将纳米孔转化为用于各种基因和表观遗传检测的超灵敏单分子平台。然而，与包括聚合酶链反应（PCR）在内的现有方法相比，低通量限制了纳米孔在生物学研究和临床环境中的应用，而生物学研究和临床环境通常需要同时检测多种生物标志物以进行准确的疾病诊断。在此，我们报告了一种用于在单个纳米孔中进行多种核酸检测的条形码探针方法。我们不是直接在纳米孔中识别不同的靶标，而是设计了一系列条形码探针来编码不同的靶标。当探针与靶标结合时，通过点击化学连接在探针上的条形码基团聚乙二醇可以特异性地调节纳米孔离子流。产生的特征信号作为编码靶标的标记。因此，在电流记录中对不同的特征信号进行计数，可以在单个纳米孔中同时分析多个靶标。通过使用一组癌症衍生的微小RNA作为靶标（一种用于癌症检测的生物标志物类型）验证了该方法的原理。
