SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12

Autor: Alfredo Garcia-Venzor, Bertha Rueda-Zarazua, Eduardo Marquez-Garcia, Vilma Maldonado, Angelica Moncada-Morales, Hiram Olivera, Irma Lopez, Joaquin Zuñiga, Jorge Melendez-Zajgla
Jazyk: angličtina
Rok vydání: 2021
Předmět:
Zdroj: Frontiers in Medicine, Vol 8 (2021)
Druh dokumentu: article
ISSN: 2296-858X
DOI: 10.3389/fmed.2021.627679
Popis: As to date, more than 49 million confirmed cases of Coronavirus Disease 19 (COVID-19) have been reported worldwide. Current diagnostic protocols use qRT-PCR for viral RNA detection, which is expensive and requires sophisticated equipment, trained personnel and previous RNA extraction. For this reason, we need a faster, direct and more versatile detection method for better epidemiological management of the COVID-19 outbreak. In this work, we propose a direct method without RNA extraction, based on the Loop-mediated isothermal amplification (LAMP) and Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR associated protein (CRISPR-Cas12) technique that allows the fast detection of SARS-CoV-2 from patient samples with high sensitivity and specificity. We obtained a limit of detection of 16 copies/μL with high specificity and at an affordable cost. The diagnostic test readout can be done with a real-time PCR thermocycler or with the naked eye in a blue-light transilluminator. Our method has been evaluated on a small set of clinical samples with promising results.
Databáze: Directory of Open Access Journals