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A Spinach molecular beacon triggered by strand displacement
We have re-engineered the fluorescent RNA aptamer Spinach to be activated in a sequence-dependent manner. The original Spinach aptamer was extended at its 5′- and 3′-ends to create Spinach.ST, which is predicted to fold into an inactive conformation and thus prevent association with the small molecu...
Autores principales: | , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Cold Spring Harbor Laboratory Press
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4105745/ https://www.ncbi.nlm.nih.gov/pubmed/24942625 http://dx.doi.org/10.1261/rna.045047.114 |
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author | Bhadra, Sanchita Ellington, Andrew D. |
author_facet | Bhadra, Sanchita Ellington, Andrew D. |
author_sort | Bhadra, Sanchita |
collection | PubMed |
description | We have re-engineered the fluorescent RNA aptamer Spinach to be activated in a sequence-dependent manner. The original Spinach aptamer was extended at its 5′- and 3′-ends to create Spinach.ST, which is predicted to fold into an inactive conformation and thus prevent association with the small molecule fluorophore DFHBI. Hybridization of a specific trigger oligonucleotide to a designed toehold leads to toehold-initiated strand displacement and refolds Spinach into the active, fluorophore-binding conformation. Spinach.ST not only specifically detects its target oligonucleotide but can discriminate readily against single-nucleotide mismatches. RNA amplicons produced during nucleic acid sequence-based amplification (NASBA) of DNA or RNA targets could be specifically detected and reported in real-time by conformational activation of Spinach.ST generated by in vitro transcription. In order to adapt any target sequence to detection by a Spinach reporter we used a primer design technique that brings together otherwise distal toehold sequences via hairpin formation. The same techniques could potentially be used to adapt common Spinach reporters to non-nucleic acid analytes, rather than by making fusions between aptamers and Spinach. |
format | Online Article Text |
id | pubmed-4105745 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Cold Spring Harbor Laboratory Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-41057452015-08-01 A Spinach molecular beacon triggered by strand displacement Bhadra, Sanchita Ellington, Andrew D. RNA Report We have re-engineered the fluorescent RNA aptamer Spinach to be activated in a sequence-dependent manner. The original Spinach aptamer was extended at its 5′- and 3′-ends to create Spinach.ST, which is predicted to fold into an inactive conformation and thus prevent association with the small molecule fluorophore DFHBI. Hybridization of a specific trigger oligonucleotide to a designed toehold leads to toehold-initiated strand displacement and refolds Spinach into the active, fluorophore-binding conformation. Spinach.ST not only specifically detects its target oligonucleotide but can discriminate readily against single-nucleotide mismatches. RNA amplicons produced during nucleic acid sequence-based amplification (NASBA) of DNA or RNA targets could be specifically detected and reported in real-time by conformational activation of Spinach.ST generated by in vitro transcription. In order to adapt any target sequence to detection by a Spinach reporter we used a primer design technique that brings together otherwise distal toehold sequences via hairpin formation. The same techniques could potentially be used to adapt common Spinach reporters to non-nucleic acid analytes, rather than by making fusions between aptamers and Spinach. Cold Spring Harbor Laboratory Press 2014-08 /pmc/articles/PMC4105745/ /pubmed/24942625 http://dx.doi.org/10.1261/rna.045047.114 Text en © 2014 Bhadra and Ellington; Published by Cold Spring Harbor Laboratory Press for the RNA Society http://creativecommons.org/licenses/by-nc/4.0/ This article is distributed exclusively by the RNA Society for the first 12 months after the full-issue publication date (see http://rnajournal.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/. |
spellingShingle | Report Bhadra, Sanchita Ellington, Andrew D. A Spinach molecular beacon triggered by strand displacement |
title | A Spinach molecular beacon triggered by strand displacement |
title_full | A Spinach molecular beacon triggered by strand displacement |
title_fullStr | A Spinach molecular beacon triggered by strand displacement |
title_full_unstemmed | A Spinach molecular beacon triggered by strand displacement |
title_short | A Spinach molecular beacon triggered by strand displacement |
title_sort | spinach molecular beacon triggered by strand displacement |
topic | Report |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4105745/ https://www.ncbi.nlm.nih.gov/pubmed/24942625 http://dx.doi.org/10.1261/rna.045047.114 |
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