Optimization and Benchmarking of RT-LAMP-CRISPR-Cas12a for the Detection of SARS-CoV-2 in Saliva.

ABSTRACT: Resource-limited settings and supply chain difficulties faced throughout the COVID-19 pandemic prompted the development of rapid and alternative methods of detecting SARS-CoV-2. These methods include reverse-transcription loop-mediated isothermal amplification (RT-LAMP), reverse-transcription recombinase polymerase amplification (RT-RPA), and CRISPR-Cas12a fluorescence detection. We describe RT-LAMP, RT-RPA, and CRISPR-Cas12a assays for the detection of the N and E-gene amplicons of SARS-CoV-2 and the optimization of various assay components, including incubation temperatures, Cas12a enzymes, reporter molecules, and the use of a lyophilized RT-LAMP master mix. We also describe the testing of a one-tube RT-LAMP-CRISPR-Cas12a assay. The one-tube assay showed promise in reducing hands-on time

SUBMITTER: Lynch CRH 

PROVIDER: S-EPMC11899638 | biostudies-literature | 2025 Feb

REPOSITORIES: biostudies-literature