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

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 reaso...

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Autores principales: Garcia-Venzor, Alfredo, Rueda-Zarazua, Bertha, Marquez-Garcia, Eduardo, Maldonado, Vilma, Moncada-Morales, Angelica, Olivera, Hiram, Lopez, Irma, Zuñiga, Joaquin, Melendez-Zajgla, Jorge
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7928313/
https://www.ncbi.nlm.nih.gov/pubmed/33681254
http://dx.doi.org/10.3389/fmed.2021.627679
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author Garcia-Venzor, Alfredo
Rueda-Zarazua, Bertha
Marquez-Garcia, Eduardo
Maldonado, Vilma
Moncada-Morales, Angelica
Olivera, Hiram
Lopez, Irma
Zuñiga, Joaquin
Melendez-Zajgla, Jorge
author_facet Garcia-Venzor, Alfredo
Rueda-Zarazua, Bertha
Marquez-Garcia, Eduardo
Maldonado, Vilma
Moncada-Morales, Angelica
Olivera, Hiram
Lopez, Irma
Zuñiga, Joaquin
Melendez-Zajgla, Jorge
author_sort Garcia-Venzor, Alfredo
collection PubMed
description 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.
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spelling pubmed-79283132021-03-04 SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12 Garcia-Venzor, Alfredo Rueda-Zarazua, Bertha Marquez-Garcia, Eduardo Maldonado, Vilma Moncada-Morales, Angelica Olivera, Hiram Lopez, Irma Zuñiga, Joaquin Melendez-Zajgla, Jorge Front Med (Lausanne) Medicine 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. Frontiers Media S.A. 2021-02-17 /pmc/articles/PMC7928313/ /pubmed/33681254 http://dx.doi.org/10.3389/fmed.2021.627679 Text en Copyright © 2021 Garcia-Venzor, Rueda-Zarazua, Marquez-Garcia, Maldonado, Moncada-Morales, Olivera, Lopez, Zuñiga and Melendez-Zajgla. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Medicine
Garcia-Venzor, Alfredo
Rueda-Zarazua, Bertha
Marquez-Garcia, Eduardo
Maldonado, Vilma
Moncada-Morales, Angelica
Olivera, Hiram
Lopez, Irma
Zuñiga, Joaquin
Melendez-Zajgla, Jorge
SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title_full SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title_fullStr SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title_full_unstemmed SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title_short SARS-CoV-2 Direct Detection Without RNA Isolation With Loop-Mediated Isothermal Amplification (LAMP) and CRISPR-Cas12
title_sort sars-cov-2 direct detection without rna isolation with loop-mediated isothermal amplification (lamp) and crispr-cas12
topic Medicine
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7928313/
https://www.ncbi.nlm.nih.gov/pubmed/33681254
http://dx.doi.org/10.3389/fmed.2021.627679
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