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Translational Enhancer Based Amplification of Toehold Sensors (TacToe) for Improved Sensitivity and Speed of Viral RNA Detection
Indexado
WoS WOS:001498877300001
Scopus SCOPUS_ID:105006911555
DOI 10.1021/ACSSYNBIO.4C00861
Año 2025
Tipo artículo de investigación

Citas Totales

Autores Afiliación Chile

Instituciones Chile

% Participación
Internacional

Autores
Afiliación Extranjera

Instituciones
Extranjeras


Abstract



Toehold switches are RNA riboswitches activated by complementary nucleic acid sequences that have shown promise as point-of-care (PoC) molecular diagnostics. However, typical implementations require an additional nucleic-acid-sequence-specific amplification step. Here, we describe a novel toehold switch with a T7-g10 translational enhancer (Tac) that amplifies the expression of the reporter gene regulated by toehold (Toe) sensors. We compare such a TacToe sensor to a previously developed sensor for detecting short RNA sequences from the Zika virus (ZIKV). We demonstrate that this one-step TacToe sensor in a transcription-translation (TxTl) reaction has an improved sensitivity to RNA and a faster time of detection, compared to the conventional toehold. Replacing the ZIVK sequence with a segment of the N-gene of SARS-nCoV, we demonstrate up to a picomolar sensitivity. Qualitatively comparable results are observed in Escherichia coli cell lysate-based homemade cell free extract (CFE), demonstrating the robustness and utility of these sensors in low-resource settings.

Revista



Revista ISSN
Acs Synthetic Biology 2161-5063

Métricas Externas



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Disciplinas de Investigación



WOS
Biochemical Research Methods
Scopus
Sin Disciplinas
SciELO
Sin Disciplinas

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Publicaciones WoS (Ediciones: ISSHP, ISTP, AHCI, SSCI, SCI), Scopus, SciELO Chile.

Colaboración Institucional



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Autores - Afiliación



Ord. Autor Género Institución - País
1 Kale, Tanvi - IISER Pune - India
Indian Institute of Science Education and Research Pune - India
2 Pednekar, Rudvi - IISER Pune - India
Indian Institute of Science Education and Research Pune - India
3 Cazaux, Severine Marianne - Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile
Pontificia Universidad Católica de Chile - Chile
Instituto Milenio de Biología Integrativa - Chile
4 Letelier, Valentina Ferrando - Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile
Pontificia Universidad Católica de Chile - Chile
Instituto Milenio de Biología Integrativa - Chile
5 Vigar, Justin R. J. - UNIV TORONTO - Canadá
University of Toronto - Canadá
6 Federici, Fernan - Núcleo Milenio en Biología Sintética y Biología de Sistemas Vegetales - Chile
Pontificia Universidad Católica de Chile - Chile
Instituto Milenio de Biología Integrativa - Chile
7 Pardee, Keith - UNIV TORONTO - Canadá
University of Toronto - Canadá
8 Athale, Chaitanya A. - IISER Pune - India
Indian Institute of Science Education and Research Pune - India

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Financiamiento



Fuente
FONDECYT
Fondo Nacional de Desarrollo Científico y Tecnológico
Fondo de Desarrollo Científico y Tecnológico
Fondo de Desarrollo Científico y Tecnológico (Fondecyt)
University of Toronto
International Development Research Centre
Canada's International Development Research Centre (IDRC)
ANID
National Agency for Research and Development
Ontario Graduate Scholarship program
Agenția Națională pentru Cercetare și Dezvoltare
University of Toronto's Emerging & Pandemic Infections Consortium
Shastri Indo-Canadian Institute (SICI)
Centre for Pharmaceutical Oncology
Shastri Indo-Canadian Institute
IISER Pune
IISER

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Agradecimientos



Agradecimiento
TK is supported by a PhD fellowship from IISER Pune. JV is supported by the Ontario Graduate Scholarship program, and awards from the University of Toronto's Emerging & Pandemic Infections Consortium and the Centre for Pharmaceutical Oncology. The work was supported by funding from the Shastri Indo-Canadian Institute (SICI) Shastri Covid-19 Pandemic Response Grant (SCPRG) and Canada's International Development Research Centre (IDRC) (109434-001) through the Canadian 2019 Novel Coronavirus (COVID-19) Rapid Research Funding Opportunity and the National Agency for Research and Development (ANID) through the ANID Millennium Science Initiative Program (ICN17_022) and Fondo de Desarrollo Cientifico y Tecnologico (FONDECYT Regular 1241452 and FONDECYT Exploracion 13220075 awarded to FF). ZIKV Sensor27B LacZ (Addgene plasmid #75006) and ZIKV NASBA 27B (Addgene plasmid #75010) were a gifts from James Collins & Alexander Green. We are grateful to Mamatharani D.V. and Krishanpal Karmodiya for help with in vitro transcription of ZIKV and Cov2 RNA from plasmid DNA. We are grateful to Sunish Radhakrishnan for discussions.
TK is supported by a PhD fellowship from IISER Pune. JV is supported by the Ontario Graduate Scholarship program, and awards from the University of Toronto\u2019s Emerging & Pandemic Infections Consortium and the Centre for Pharmaceutical Oncology. The work was supported by funding from the Shastri Indo-Canadian Institute (SICI) Shastri Covid-19 Pandemic Response Grant (SCPRG) and Canada\u2019s International Development Research Centre (IDRC) (109434-001) through the Canadian 2019 Novel Coronavirus (COVID-19) Rapid Research Funding Opportunity and the National Agency for Research and Development (ANID) through the ANID Millennium Science Initiative Program (ICN17_022) and Fondo de Desarrollo Cienti\u0301fico y Tecnolo\u0301gico (FONDECYT Regular 1241452 and FONDECYT Exploracio\u0301n 13220075 awarded to FF). ZIKV Sensor27B LacZ (Addgene plasmid #75006) and ZIKV NASBA 27B (Addgene plasmid #75010) were a gifts from James Collins & Alexander Green. We are grateful to Mamatharani D.V. and Krishanpal Karmodiya for help with in vitro transcription of ZIKV and Cov2 RNA from plasmid DNA. We are grateful to Sunish Radhakrishnan for discussions.

Muestra la fuente de financiamiento declarada en la publicación.