Department of Chemistry, Jeonbuk National University, Jeonju, 54896, South Korea.
Department of Laboratory Medicine, Jeonbuk National University Medical School and Hospital, Jeonju, 54896, South Korea.
Talanta. 2023 Jan 15;252:123835. doi: 10.1016/j.talanta.2022.123835. Epub 2022 Aug 14.
In this paper we present a new method for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), targeting a specific region "N gene." Under isothermal reaction conditions, we integrated ligation (Lig; high selectivity) and recombinase polymerase amplification (RPA; high sensitivity) processes, obtaining a robust method of detection. For point-of-care testing, we incorporated our laboratory-produced pyrophosphate ion (PPi)-sensing probe (PK-probe) for colorimetric analysis of the reaction. The total detection system was efficient and effective at diagnosing this RNA virus-mediated disease rapidly (30 min). In a full-genome SARS-CoV-2 study, our PK-probe/Lig-RPA system functioned with a limit of detection of 1160 copies/ml, with a single-mismatch level of selectively, and it was highly selective even in the presence of bacterial genomes commonly found in the human mouth and nose. This robust, straightforward, selective, efficient, and ultrasensitive colorimetric detection method, with potential for point-of-care analysis, should also be effective in detecting a diverse range of other RNA-based diseases.
本文提出了一种新的方法来检测严重急性呼吸综合征冠状病毒 2（SARS-CoV-2），该方法针对特定区域“N 基因”。在等温反应条件下，我们整合了连接（Lig；高选择性）和重组酶聚合酶扩增（RPA；高灵敏度）过程，获得了一种强大的检测方法。为了进行即时检测，我们结合了实验室生产的焦磷酸离子（PPi）感应探针（PK-probe）进行反应的比色分析。该总检测系统在快速（30 分钟）诊断这种 RNA 病毒介导的疾病方面非常有效。在全基因组 SARS-CoV-2 研究中，我们的 PK-probe/Lig-RPA 系统具有 1160 拷贝/ml 的检测限，具有单碱基错配水平的选择性，即使在常见于人体口腔和鼻腔的细菌基因组存在的情况下，也具有高度选择性。这种强大、简单、选择性、高效、超灵敏的比色检测方法，具有即时分析的潜力，也应该能够有效检测多种其他基于 RNA 的疾病。
