Department of Biomedical Engineering, University of Connecticut, Storrs, CT 06269, USA.
Institute of Materials Science, University of Connecticut, Storrs, CT 06269, USA.
Angew Chem Int Ed Engl. 2022 Aug 8;61(32):e202203826. doi: 10.1002/anie.202203826. Epub 2022 May 31.
The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems have recently received notable attention for their applications in nucleic acid detection. Despite many attempts, the majority of current CRISPR-based biosensors in infectious respiratory disease diagnostic applications still require target preamplifications. This study reports a new biosensor for amplification-free nucleic acid detection via harnessing the trans-cleavage mechanism of Cas13a and ultrasensitive graphene field-effect transistors (gFETs). CRISPR Cas13a-gFET achieves the detection of SARS-CoV-2 and respiratory syncytial virus (RSV) genome down to 1 attomolar without target preamplifications. Additionally, we validate the detection performance using clinical SARS-CoV-2 samples, including those with low viral loads (Ct value >30). Overall, these findings establish our CRISPR Cas13a-gFET among the most sensitive amplification-free nucleic acid diagnostic platforms to date.
簇状规律间隔短回文重复 (CRISPR)/CRISPR 相关 (Cas) 系统因其在核酸检测中的应用而受到广泛关注。尽管进行了许多尝试，但目前大多数基于 CRISPR 的生物传感器在传染性呼吸道疾病诊断应用中仍需要靶标预扩增。本研究报告了一种新的生物传感器，用于通过利用 Cas13a 的转切割机制和超灵敏石墨烯场效应晶体管 (gFET) 进行无扩增的核酸检测。CRISPR Cas13a-gFET 可在无需靶标预扩增的情况下，检测到低至 1 飞摩尔的 SARS-CoV-2 和呼吸道合胞病毒 (RSV) 基因组。此外，我们使用临床 SARS-CoV-2 样本验证了检测性能，包括那些病毒载量较低 (Ct 值 >30) 的样本。总的来说，这些发现确立了我们的 CRISPR Cas13a-gFET 是迄今为止最灵敏的无扩增核酸诊断平台之一。
