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Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing

The sensitivity of FRET-based sensing is usually limited by the spectral overlaps of the FRET donor and acceptor, which generate a poor signal-to-noise ratio. To overcome this limitation, a quenched donor presenting a large Stokes shift can be combined with a bright acceptor to perform Dark Resonanc...

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Autores principales: Barnoin, Guillaume, Shaya, Janah, Richert, Ludovic, Le, Hoang-Ngoan, Vincent, Steve, Guérineau, Vincent, Mély, Yves, Michel, Benoît Y, Burger, Alain
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8266640/
https://www.ncbi.nlm.nih.gov/pubmed/33872373
http://dx.doi.org/10.1093/nar/gkab237
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author Barnoin, Guillaume
Shaya, Janah
Richert, Ludovic
Le, Hoang-Ngoan
Vincent, Steve
Guérineau, Vincent
Mély, Yves
Michel, Benoît Y
Burger, Alain
author_facet Barnoin, Guillaume
Shaya, Janah
Richert, Ludovic
Le, Hoang-Ngoan
Vincent, Steve
Guérineau, Vincent
Mély, Yves
Michel, Benoît Y
Burger, Alain
author_sort Barnoin, Guillaume
collection PubMed
description The sensitivity of FRET-based sensing is usually limited by the spectral overlaps of the FRET donor and acceptor, which generate a poor signal-to-noise ratio. To overcome this limitation, a quenched donor presenting a large Stokes shift can be combined with a bright acceptor to perform Dark Resonance Energy Transfer (DRET). The consequent fluorogenic response from the acceptor considerably improves the signal-to-noise ratio. To date, DRET has mainly relied on a donor that is covalently bound to the acceptor. In this context, our aim was to develop the first intermolecular DRET pair for specific sensing of nucleic acid sequences. To this end, we designed DFK, a push–pull probe based on a fluorenyl π-platform that is strongly quenched in water. DFK was incorporated into a series of oligonucleotides and used as a DRET donor with Cy5-labeled complementary sequences. In line with our expectations, excitation of the dark donor in the double-labeled duplex switched on the far-red Cy5 emission and remained free of cross-excitation. The DRET mechanism was supported by time-resolved fluorescence measurements. This concept was then applied with binary probes, which confirmed the distance dependence of DRET as well as its potency in detecting sequences of interest with low background noise.
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spelling pubmed-82666402021-07-09 Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing Barnoin, Guillaume Shaya, Janah Richert, Ludovic Le, Hoang-Ngoan Vincent, Steve Guérineau, Vincent Mély, Yves Michel, Benoît Y Burger, Alain Nucleic Acids Res Methods Online The sensitivity of FRET-based sensing is usually limited by the spectral overlaps of the FRET donor and acceptor, which generate a poor signal-to-noise ratio. To overcome this limitation, a quenched donor presenting a large Stokes shift can be combined with a bright acceptor to perform Dark Resonance Energy Transfer (DRET). The consequent fluorogenic response from the acceptor considerably improves the signal-to-noise ratio. To date, DRET has mainly relied on a donor that is covalently bound to the acceptor. In this context, our aim was to develop the first intermolecular DRET pair for specific sensing of nucleic acid sequences. To this end, we designed DFK, a push–pull probe based on a fluorenyl π-platform that is strongly quenched in water. DFK was incorporated into a series of oligonucleotides and used as a DRET donor with Cy5-labeled complementary sequences. In line with our expectations, excitation of the dark donor in the double-labeled duplex switched on the far-red Cy5 emission and remained free of cross-excitation. The DRET mechanism was supported by time-resolved fluorescence measurements. This concept was then applied with binary probes, which confirmed the distance dependence of DRET as well as its potency in detecting sequences of interest with low background noise. Oxford University Press 2021-04-19 /pmc/articles/PMC8266640/ /pubmed/33872373 http://dx.doi.org/10.1093/nar/gkab237 Text en © The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/ (https://creativecommons.org/licenses/by-nc/4.0/) ), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Methods Online
Barnoin, Guillaume
Shaya, Janah
Richert, Ludovic
Le, Hoang-Ngoan
Vincent, Steve
Guérineau, Vincent
Mély, Yves
Michel, Benoît Y
Burger, Alain
Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title_full Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title_fullStr Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title_full_unstemmed Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title_short Intermolecular dark resonance energy transfer (DRET): upgrading fluorogenic DNA sensing
title_sort intermolecular dark resonance energy transfer (dret): upgrading fluorogenic dna sensing
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8266640/
https://www.ncbi.nlm.nih.gov/pubmed/33872373
http://dx.doi.org/10.1093/nar/gkab237
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