Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides

Detalhes bibliográficos
Autor(a) principal: Ortiz, Mayreli
Data de Publicação: 2021
Outros Autores: Jauset-Rubio, Miriam, Skouridou, Vasso, Machado, Diana, Viveiros, Miguel, Clark, Taane G., Simonova, Anna, Kodr, David, Hocek, Michal, O'Sullivan, Ciara K.
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10362/130385
Resumo: Funding Information: This project received partial funding from the European Union’s Horizon 2020 Research and Innovation Programme under grant agreement No 767325, the Czech Science Foundation (20-00885X to M.H.), and the Fundação para a Ciência e a Tecnologia, Portugal (Grant - UID/Multi/04413/2020). The authors thank Fraunhofer ICT-IMM for the provision of gold electrode arrays. The Ph.D. scholarship of D. Kodr from the Department of Chemistry of Natural Compounds of the University of Chemistry and Technology Prague is also acknowledged. Publisher Copyright: © 2021 The Authors. Published by American Chemical Society.
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spelling Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotidesferrocene-labeled nucleotidesKlenow (exo-) DNA polymerasenucleoside triphosphatessingle-nucleotide polymorphism (SNP)single-point mutationsolid-phase primer elongationBioengineeringInstrumentationProcess Chemistry and TechnologyFluid Flow and Transfer ProcessesInfectious DiseasesSDG 3 - Good Health and Well-beingSDG 9 - Industry, Innovation, and InfrastructureSDG 10 - Reduced InequalitiesSDG 12 - Responsible Consumption and ProductionSDG 17 - Partnerships for the GoalsFunding Information: This project received partial funding from the European Union’s Horizon 2020 Research and Innovation Programme under grant agreement No 767325, the Czech Science Foundation (20-00885X to M.H.), and the Fundação para a Ciência e a Tecnologia, Portugal (Grant - UID/Multi/04413/2020). The authors thank Fraunhofer ICT-IMM for the provision of gold electrode arrays. The Ph.D. scholarship of D. Kodr from the Department of Chemistry of Natural Compounds of the University of Chemistry and Technology Prague is also acknowledged. Publisher Copyright: © 2021 The Authors. Published by American Chemical Society.Here, we report the electrochemical detection of single-point mutations using solid-phase isothermal primer elongation with redox-labeled oligonucleotides. A single-base mutation associated with resistance to rifampicin, an antibiotic commonly used for the treatment of Mycobacterium tuberculosis, was used as a model system to demonstrate a proof-of-concept of the approach. Four 5′-thiolated primers, designed to be complementary with the same fragment of the target sequence and differing only in the last base, addressing the polymorphic site, were self-assembled via chemisorption on individual gold electrodes of an array. Following hybridization with single-stranded DNA, Klenow (exo-) DNA polymerase-mediated primer extension with ferrocene-labeled 2′-deoxyribonucleoside triphosphates (dNFcTPs) was only observed to proceed at the electrode where there was full complementarity between the surface-tethered probe and the target DNA being interrogated. We tested all four ferrocenylethynyl-linked dNTPs and optimized the ratio of labeled/natural nucleotides to achieve maximum sensitivity. Following a 20 min hybridization step, Klenow (exo-) DNA polymerase-mediated primer elongation at 37 °C for 5 min was optimal for the enzymatic incorporation of a ferrocene-labeled nucleotide, achieving unequivocal electrochemical detection of a single-point mutation in 14 samples of genomic DNA extracted from Mycobacterium tuberculosis strains. The approach is rapid, cost-effective, facile, and can be extended to multiplexed electrochemical single-point mutation genotyping.TB, HIV and opportunistic diseases and pathogens (THOP)Global Health and Tropical Medicine (GHTM)Instituto de Higiene e Medicina Tropical (IHMT)RUNOrtiz, MayreliJauset-Rubio, MiriamSkouridou, VassoMachado, DianaViveiros, MiguelClark, Taane G.Simonova, AnnaKodr, DavidHocek, MichalO'Sullivan, Ciara K.2023-04-22T00:31:45Z2021-12-242021-12-24T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/article10application/pdfhttp://hdl.handle.net/10362/130385eng2379-3694PURE: 35225426https://doi.org/10.1021/acssensors.1c01710info: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:RCAAP2024-03-11T05:09:03Zoai:run.unl.pt:10362/130385Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:46:45.573247Repositório 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 Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
title Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
spellingShingle Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
Ortiz, Mayreli
ferrocene-labeled nucleotides
Klenow (exo-) DNA polymerase
nucleoside triphosphates
single-nucleotide polymorphism (SNP)
single-point mutation
solid-phase primer elongation
Bioengineering
Instrumentation
Process Chemistry and Technology
Fluid Flow and Transfer Processes
Infectious Diseases
SDG 3 - Good Health and Well-being
SDG 9 - Industry, Innovation, and Infrastructure
SDG 10 - Reduced Inequalities
SDG 12 - Responsible Consumption and Production
SDG 17 - Partnerships for the Goals
title_short Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
title_full Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
title_fullStr Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
title_full_unstemmed Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
title_sort Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
author Ortiz, Mayreli
author_facet Ortiz, Mayreli
Jauset-Rubio, Miriam
Skouridou, Vasso
Machado, Diana
Viveiros, Miguel
Clark, Taane G.
Simonova, Anna
Kodr, David
Hocek, Michal
O'Sullivan, Ciara K.
author_role author
author2 Jauset-Rubio, Miriam
Skouridou, Vasso
Machado, Diana
Viveiros, Miguel
Clark, Taane G.
Simonova, Anna
Kodr, David
Hocek, Michal
O'Sullivan, Ciara K.
author2_role author
author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv TB, HIV and opportunistic diseases and pathogens (THOP)
Global Health and Tropical Medicine (GHTM)
Instituto de Higiene e Medicina Tropical (IHMT)
RUN
dc.contributor.author.fl_str_mv Ortiz, Mayreli
Jauset-Rubio, Miriam
Skouridou, Vasso
Machado, Diana
Viveiros, Miguel
Clark, Taane G.
Simonova, Anna
Kodr, David
Hocek, Michal
O'Sullivan, Ciara K.
dc.subject.por.fl_str_mv ferrocene-labeled nucleotides
Klenow (exo-) DNA polymerase
nucleoside triphosphates
single-nucleotide polymorphism (SNP)
single-point mutation
solid-phase primer elongation
Bioengineering
Instrumentation
Process Chemistry and Technology
Fluid Flow and Transfer Processes
Infectious Diseases
SDG 3 - Good Health and Well-being
SDG 9 - Industry, Innovation, and Infrastructure
SDG 10 - Reduced Inequalities
SDG 12 - Responsible Consumption and Production
SDG 17 - Partnerships for the Goals
topic ferrocene-labeled nucleotides
Klenow (exo-) DNA polymerase
nucleoside triphosphates
single-nucleotide polymorphism (SNP)
single-point mutation
solid-phase primer elongation
Bioengineering
Instrumentation
Process Chemistry and Technology
Fluid Flow and Transfer Processes
Infectious Diseases
SDG 3 - Good Health and Well-being
SDG 9 - Industry, Innovation, and Infrastructure
SDG 10 - Reduced Inequalities
SDG 12 - Responsible Consumption and Production
SDG 17 - Partnerships for the Goals
description Funding Information: This project received partial funding from the European Union’s Horizon 2020 Research and Innovation Programme under grant agreement No 767325, the Czech Science Foundation (20-00885X to M.H.), and the Fundação para a Ciência e a Tecnologia, Portugal (Grant - UID/Multi/04413/2020). The authors thank Fraunhofer ICT-IMM for the provision of gold electrode arrays. The Ph.D. scholarship of D. Kodr from the Department of Chemistry of Natural Compounds of the University of Chemistry and Technology Prague is also acknowledged. Publisher Copyright: © 2021 The Authors. Published by American Chemical Society.
publishDate 2021
dc.date.none.fl_str_mv 2021-12-24
2021-12-24T00:00:00Z
2023-04-22T00:31:45Z
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/10362/130385
url http://hdl.handle.net/10362/130385
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv 2379-3694
PURE: 35225426
https://doi.org/10.1021/acssensors.1c01710
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv 10
application/pdf
dc.source.none.fl_str_mv reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
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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
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