Department of Respiratory and Critical Care Medicine, West China Medical School/West China Hospital, Sichuan University, Chengdu 610041, China.
College of Biomass Science and Engineering, Healthy Food Evaluation Research Center, Sichuan University, Chengdu 610065, China.
Anal Chem. 2021 Feb 23;93(7):3393-3402. doi: 10.1021/acs.analchem.0c04303. Epub 2021 Jan 29.
The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global health emergency, and its gene mutation and evolution further posed uncertainty of epidemic risk. Herein, we reported a light-up CRISPR-Cas13 transcription amplification method, which enables the detection of SARS-CoV-2 and its mutated variants. Sequence specificity was ensured by both the ligation process and Cas13a/crRNA recognition, allowing us to identify viral RNA mutation. Light-up RNA aptamer allows sensitive output of amplification signals via target-activated ribonuclease activity of CRISPR-Cas13a. The RNA virus assay has been designed to detect coronavirus, SARS-CoV-2, Middle East respiratory syndrome (MERS), and SARS, as well as the influenza viruses such as, H1N1, H7N9, and H9N2. It was accommodated to sense as low as 82 copies of SARS-CoV-2. Particularly, it allowed us to strictly discriminate key mutation of the SARS-CoV-2 variant, D614G, which may induce higher epidemic and pathogenetic risk. The proposed RNA virus assays are promising for point-of-care monitoring of SARS-CoV-2 and its risking variants.
严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 的爆发引发了全球卫生紧急事件，其基因变异和进化进一步带来了疫情风险的不确定性。在此，我们报告了一种点亮型 CRISPR-Cas13 转录扩增方法，该方法可用于检测 SARS-CoV-2 及其突变变体。通过连接过程和 Cas13a/crRNA 识别确保了序列特异性，使我们能够识别病毒 RNA 突变。点亮型 RNA 适体通过 CRISPR-Cas13a 的靶激活核糖核酸酶活性实现了扩增信号的灵敏输出。该 RNA 病毒检测已被设计用于检测冠状病毒、SARS-CoV-2、中东呼吸综合征 (MERS) 和 SARS，以及流感病毒，如 H1N1、H7N9 和 H9N2。它可以检测到低至 82 拷贝的 SARS-CoV-2。特别是，它允许我们严格区分 SARS-CoV-2 变体的关键突变 D614G，这可能会导致更高的流行和发病风险。所提出的 RNA 病毒检测方法有望用于 SARS-CoV-2 及其风险变体的即时护理监测。
