Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot

Gene sequencing in back of reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is the current approach for discriminating infections produced by different severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants in the clinic. However, sequencing is often a time-consu...

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Autores: Ruiz, Raúl, Montagud-Martínez, Roser, Dorta-Gorrín, Alexis, Pablo-Marcos, Daniel, Gozalo, Mónica, Calvo-Montes, Jorge, Navas, Jesús, Rodrigo, Guillermo
Tipo de recurso: artículo
Estado:Versión publicada
Fecha de publicación:2024
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/361532
Acceso en línea:http://hdl.handle.net/10261/361532
https://api.elsevier.com/content/abstract/scopus_id/85190167695
Access Level:acceso abierto
Palabra clave:Fluorescence
SARS-CoV-2
Genetics
Probes
Infectious diseases
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spelling Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR potRuiz, RaúlMontagud-Martínez, RoserDorta-Gorrín, AlexisPablo-Marcos, DanielGozalo, MónicaCalvo-Montes, JorgeNavas, JesúsRodrigo, GuillermoFluorescenceSARS-CoV-2GeneticsProbesInfectious diseasesGene sequencing in back of reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is the current approach for discriminating infections produced by different severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants in the clinic. However, sequencing is often a time-consuming step, which hinders the deployment of a very fast response during a pandemic. Here, we propose to run a CRISPR-Cas12a reaction after completing the RT-qPCR and in the very same pot to detect with high specificity genetic marks characterizing variants of concern. A crRNA was appropriately designed to detect the S gene of the SARS-CoV-2 Omicron BA.1 variant. A significant response with >20-fold dynamic range was obtained for the Omicron BA.1 S gene, while the Delta S gene did not produce any detectable signal. The sensitivity of the method was analyzed with a series of diluted samples and different Cas12a nucleases. A correlation between the RT-qPCR CT values and the CRISPR-Cas12a reaction signals was observed. Variant discrimination with the CRISPR-Cas12a reaction was possible in some minutes with high accuracy from patient samples. In conclusion, CRISPR-Cas systems seem ready to be exploited in the clinic to boost personalized diagnoses and accelerate epidemiological surveillance in a cost-effective way.Work supported by Generalitat Valenciana (GVA-COVID19/2021/036), Ministerio de Ciencia e Innovación (PDC2022−133941-I00; cofunded by NextGenerationEU), CRUE andBanco Santander (Fondo Supera Covid-19), CSIC PTI SaludGlobal (NextGenerationEU Fund, regulation 2020/2094), andIDIVAL (INNVAL21/13).Peer reviewedAmerican Chemical SocietyGeneralitat ValencianaAgencia Estatal de Investigación (España)Ministerio de Ciencia e Innovación (España)Conferencia de Rectores de las Universidades EspañolasBanco SantanderCSIC - Plataforma Temática Interdisciplinar del CSIC Salud Global (PTI Salud Global)European CommissionConsejo Superior de Investigaciones Científicas (España)Ministerio de Ciencia, Innovación y Universidades (España)Rodrigo, Guillermo [0000-0002-1871-9617]Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]202420242024info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Publisher's versioninfo:eu-repo/semantics/publishedVersionapplication/pdfhttp://hdl.handle.net/10261/361532https://api.elsevier.com/content/abstract/scopus_id/85190167695reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Inglés#PLACEHOLDER_PARENT_METADATA_VALUE#info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PDC2022-133941-I00Ruiz, Raúl; Montagud-Martínez, Roser; Dorta-Gorrín, Alexis; Pablo-Marcos, Daniel; Gozalo, Mónica; Calvo-Montes, Jorge; Navas, Jesús; Rodrigo, Guillermo; 2024; Supplementary Material: Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot [Dataset]; Amercian Chemical Society; https://doi.org/10.1021/acsomega.3c09717https://doi.org/10.1021/acsomega.3c09717Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/3615322026-05-22T06:33:51Z
dc.title.none.fl_str_mv Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
title Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
spellingShingle Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
Ruiz, Raúl
Fluorescence
SARS-CoV-2
Genetics
Probes
Infectious diseases
title_short Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
title_full Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
title_fullStr Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
title_full_unstemmed Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
title_sort Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot
dc.creator.none.fl_str_mv Ruiz, Raúl
Montagud-Martínez, Roser
Dorta-Gorrín, Alexis
Pablo-Marcos, Daniel
Gozalo, Mónica
Calvo-Montes, Jorge
Navas, Jesús
Rodrigo, Guillermo
author Ruiz, Raúl
author_facet Ruiz, Raúl
Montagud-Martínez, Roser
Dorta-Gorrín, Alexis
Pablo-Marcos, Daniel
Gozalo, Mónica
Calvo-Montes, Jorge
Navas, Jesús
Rodrigo, Guillermo
author_role author
author2 Montagud-Martínez, Roser
Dorta-Gorrín, Alexis
Pablo-Marcos, Daniel
Gozalo, Mónica
Calvo-Montes, Jorge
Navas, Jesús
Rodrigo, Guillermo
author2_role author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Generalitat Valenciana
Agencia Estatal de Investigación (España)
Ministerio de Ciencia e Innovación (España)
Conferencia de Rectores de las Universidades Españolas
Banco Santander
CSIC - Plataforma Temática Interdisciplinar del CSIC Salud Global (PTI Salud Global)
European Commission
Consejo Superior de Investigaciones Científicas (España)
Ministerio de Ciencia, Innovación y Universidades (España)
Rodrigo, Guillermo [0000-0002-1871-9617]
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Fluorescence
SARS-CoV-2
Genetics
Probes
Infectious diseases
topic Fluorescence
SARS-CoV-2
Genetics
Probes
Infectious diseases
description Gene sequencing in back of reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is the current approach for discriminating infections produced by different severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants in the clinic. However, sequencing is often a time-consuming step, which hinders the deployment of a very fast response during a pandemic. Here, we propose to run a CRISPR-Cas12a reaction after completing the RT-qPCR and in the very same pot to detect with high specificity genetic marks characterizing variants of concern. A crRNA was appropriately designed to detect the S gene of the SARS-CoV-2 Omicron BA.1 variant. A significant response with >20-fold dynamic range was obtained for the Omicron BA.1 S gene, while the Delta S gene did not produce any detectable signal. The sensitivity of the method was analyzed with a series of diluted samples and different Cas12a nucleases. A correlation between the RT-qPCR CT values and the CRISPR-Cas12a reaction signals was observed. Variant discrimination with the CRISPR-Cas12a reaction was possible in some minutes with high accuracy from patient samples. In conclusion, CRISPR-Cas systems seem ready to be exploited in the clinic to boost personalized diagnoses and accelerate epidemiological surveillance in a cost-effective way.
publishDate 2024
dc.date.none.fl_str_mv 2024
2024
2024
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Publisher's version
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/361532
https://api.elsevier.com/content/abstract/scopus_id/85190167695
url http://hdl.handle.net/10261/361532
https://api.elsevier.com/content/abstract/scopus_id/85190167695
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv #PLACEHOLDER_PARENT_METADATA_VALUE#
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PDC2022-133941-I00
Ruiz, Raúl; Montagud-Martínez, Roser; Dorta-Gorrín, Alexis; Pablo-Marcos, Daniel; Gozalo, Mónica; Calvo-Montes, Jorge; Navas, Jesús; Rodrigo, Guillermo; 2024; Supplementary Material: Rapid and accurate detection of the SARS-CoV-2 Omicron variant with a CRISPR-Cas12a reaction in the RT-qPCR pot [Dataset]; Amercian Chemical Society; https://doi.org/10.1021/acsomega.3c09717
https://doi.org/10.1021/acsomega.3c09717
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv American Chemical Society
publisher.none.fl_str_mv American Chemical Society
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
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