Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection
Autor(a) principal: | |
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Data de Publicação: | 2021 |
Outros Autores: | , , , , , , , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/10400.6/11930 |
Resumo: | The fast spread of SARS-CoV-2 has led to a global pandemic, calling for fast and accurate assays to allow infection diagnosis and prevention of transmission. We aimed to develop a molecular beacon (MB)-based detection assay for SARS-CoV-2, designed to detect the ORF1ab and S genes, proposing a two-stage COVID-19 testing strategy. The novelty of this work lies in the design and optimization of two MBs for detection of SARS-CoV-2, namely, concentration, fluorescence plateaus of hybridization, reaction temperature and real-time results. We also identify putative G-quadruplex (G4) regions in the genome of SARS-CoV-2. A total of 458 nasopharyngeal and throat swab samples (426 positive and 32 negative) were tested with the MB assay and the fluorescence levels compared with the cycle threshold (Ct) values obtained from a commercial RT-PCR test in terms of test duration, sensitivity, and specificity. Our results show that the samples with higher fluorescence levels correspond to those with low Ct values, suggesting a correlation between viral load and increased MB fluorescence. The proposed assay represents a fast (total duration of 2 h 20 min including amplification and fluorescence reading stages) and simple way of detecting SARS-CoV-2 in clinical samples from the upper respiratory tract. |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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7160 |
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Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 DetectionRNA virusesSARS-CoV-2COVID-19Fluorescence detectionDiagnosisThe fast spread of SARS-CoV-2 has led to a global pandemic, calling for fast and accurate assays to allow infection diagnosis and prevention of transmission. We aimed to develop a molecular beacon (MB)-based detection assay for SARS-CoV-2, designed to detect the ORF1ab and S genes, proposing a two-stage COVID-19 testing strategy. The novelty of this work lies in the design and optimization of two MBs for detection of SARS-CoV-2, namely, concentration, fluorescence plateaus of hybridization, reaction temperature and real-time results. We also identify putative G-quadruplex (G4) regions in the genome of SARS-CoV-2. A total of 458 nasopharyngeal and throat swab samples (426 positive and 32 negative) were tested with the MB assay and the fluorescence levels compared with the cycle threshold (Ct) values obtained from a commercial RT-PCR test in terms of test duration, sensitivity, and specificity. Our results show that the samples with higher fluorescence levels correspond to those with low Ct values, suggesting a correlation between viral load and increased MB fluorescence. The proposed assay represents a fast (total duration of 2 h 20 min including amplification and fluorescence reading stages) and simple way of detecting SARS-CoV-2 in clinical samples from the upper respiratory tract.This work was supported by project No. 149 “Track and Trace COVID-19” funded in first edition “RESEARCH 4 COVID-19” by Fundação para a Ciência e a Tecnologia (FCT) in collaboration with Agência de Investigação Clínica e Inovação Biomédica (AICIB)uBibliorumCarvalho, JosuéNunes, J. LopesFigueiredo, JoanaSantos, TiagoMiranda, AndréRiscado, MicaelaSousa, FaniDuarte, A. P.Socorro, SílviaTomaz, CândidaFelgueiras, MafaldaTeixeira, RuiFaria, ConceiçãoCruz, Carla2022-01-20T15:35:49Z2021-102021-10-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10400.6/11930eng10.3390/s21217015info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-12-15T09:54:42Zoai:ubibliorum.ubi.pt:10400.6/11930Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T00:51:36.459060Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
title |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
spellingShingle |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection Carvalho, Josué RNA viruses SARS-CoV-2 COVID-19 Fluorescence detection Diagnosis |
title_short |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
title_full |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
title_fullStr |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
title_full_unstemmed |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
title_sort |
Molecular Beacon Assay Development for Severe Acute Respiratory Syndrome Coronavirus 2 Detection |
author |
Carvalho, Josué |
author_facet |
Carvalho, Josué Nunes, J. Lopes Figueiredo, Joana Santos, Tiago Miranda, André Riscado, Micaela Sousa, Fani Duarte, A. P. Socorro, Sílvia Tomaz, Cândida Felgueiras, Mafalda Teixeira, Rui Faria, Conceição Cruz, Carla |
author_role |
author |
author2 |
Nunes, J. Lopes Figueiredo, Joana Santos, Tiago Miranda, André Riscado, Micaela Sousa, Fani Duarte, A. P. Socorro, Sílvia Tomaz, Cândida Felgueiras, Mafalda Teixeira, Rui Faria, Conceição Cruz, Carla |
author2_role |
author author author author author author author author author author author author author |
dc.contributor.none.fl_str_mv |
uBibliorum |
dc.contributor.author.fl_str_mv |
Carvalho, Josué Nunes, J. Lopes Figueiredo, Joana Santos, Tiago Miranda, André Riscado, Micaela Sousa, Fani Duarte, A. P. Socorro, Sílvia Tomaz, Cândida Felgueiras, Mafalda Teixeira, Rui Faria, Conceição Cruz, Carla |
dc.subject.por.fl_str_mv |
RNA viruses SARS-CoV-2 COVID-19 Fluorescence detection Diagnosis |
topic |
RNA viruses SARS-CoV-2 COVID-19 Fluorescence detection Diagnosis |
description |
The fast spread of SARS-CoV-2 has led to a global pandemic, calling for fast and accurate assays to allow infection diagnosis and prevention of transmission. We aimed to develop a molecular beacon (MB)-based detection assay for SARS-CoV-2, designed to detect the ORF1ab and S genes, proposing a two-stage COVID-19 testing strategy. The novelty of this work lies in the design and optimization of two MBs for detection of SARS-CoV-2, namely, concentration, fluorescence plateaus of hybridization, reaction temperature and real-time results. We also identify putative G-quadruplex (G4) regions in the genome of SARS-CoV-2. A total of 458 nasopharyngeal and throat swab samples (426 positive and 32 negative) were tested with the MB assay and the fluorescence levels compared with the cycle threshold (Ct) values obtained from a commercial RT-PCR test in terms of test duration, sensitivity, and specificity. Our results show that the samples with higher fluorescence levels correspond to those with low Ct values, suggesting a correlation between viral load and increased MB fluorescence. The proposed assay represents a fast (total duration of 2 h 20 min including amplification and fluorescence reading stages) and simple way of detecting SARS-CoV-2 in clinical samples from the upper respiratory tract. |
publishDate |
2021 |
dc.date.none.fl_str_mv |
2021-10 2021-10-01T00:00:00Z 2022-01-20T15:35:49Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/10400.6/11930 |
url |
http://hdl.handle.net/10400.6/11930 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
10.3390/s21217015 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
instname_str |
Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
instacron_str |
RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
collection |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository.name.fl_str_mv |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
repository.mail.fl_str_mv |
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1799136404779827200 |