Department of Bioengineering, Rice University, Houston, TX, USA.
Torus Biosystems, Cambridge, MA, USA.
Nat Biomed Eng. 2021 Jul;5(7):702-712. doi: 10.1038/s41551-021-00755-4. Epub 2021 Jul 1.
Assays for the molecular detection of nucleic acids are typically constrained by the level of multiplexing (this is the case for the quantitative polymerase chain reaction (qPCR) and for isothermal amplification), turnaround times (as with microarrays and next-generation sequencing), quantification accuracy (isothermal amplification, microarrays and nanopore sequencing) or specificity for single-nucleotide differences (microarrays and nanopore sequencing). Here we show that a portable and battery-powered PCR assay performed in a toroidal convection chamber housing a microarray of fluorescently quenched oligonucleotide probes allows for the rapid and sensitive quantification of multiple DNA targets with single-nucleotide discrimination. The assay offers a limit of detection of 10 DNA copies within 30 min of turnaround time and a dynamic range spanning 4 orders of magnitude of DNA concentration, and we show its performance by detecting 20 genomic loci and 30 single-nucleotide polymorphisms in human genomic DNA samples, and 15 bacterial species in clinical isolates. Portable devices for the fast and highly multiplexed detection of nucleic acids may offer advantages in point-of-care diagnostics.
用于分子检测核酸的分析方法通常受到多重性的限制（这适用于定量聚合酶链反应（qPCR）和等温扩增）、周转时间（如微阵列和下一代测序）、定量准确性（等温扩增、微阵列和纳米孔测序）或单核苷酸差异的特异性（微阵列和纳米孔测序）。在这里，我们展示了一种便携式和电池供电的 PCR 分析方法，在一个环形对流腔中进行，其中包含一个荧光猝灭寡核苷酸探针的微阵列，可以快速灵敏地定量多种 DNA 靶标，具有单核苷酸分辨能力。该分析方法的检测限为 30 分钟内的 10 个 DNA 拷贝，检测范围跨越 DNA 浓度的 4 个数量级，我们通过检测人类基因组 DNA 样本中的 20 个基因组基因座和 30 个单核苷酸多态性，以及临床分离株中的 15 种细菌，展示了其性能。用于快速和高度多重化检测核酸的便携式设备可能在即时诊断方面具有优势。
