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Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology

1
Nanoscale Science Program, Department of Chemistry, University of North Carolina at Charlotte, Charlotte, NC 28223, USA
2
Laboratory of Solution Chemistry of Advanced Materials and Technologies, ITMO University, Lomonosova St. 9, 191002 St. Petersburg, Russia
*
Author to whom correspondence should be addressed.
Academic Editor: Marina A. Dobrovolskaia
Molecules 2018, 23(12), 3178; https://doi.org/10.3390/molecules23123178
Received: 7 November 2018 / Revised: 29 November 2018 / Accepted: 29 November 2018 / Published: 2 December 2018
(This article belongs to the Special Issue Therapeutic Nucleic Acids: Past, Present, and Future)
RNA aptamers selected to bind fluorophores and activate their fluorescence offer a simple and modular way to visualize native RNAs in cells. Split aptamers which are inactive until the halves are brought within close proximity can become useful for visualizing the dynamic actions of RNA assemblies and their interactions in real time with low background noise and eliminated necessity for covalently attached dyes. Here, we design and test several sets of F30 Broccoli aptamer splits, that we call fluorets, to compare their relative fluorescence and physicochemical stabilities. We show that the splits can be simply assembled either through one-pot thermal annealing or co-transcriptionally, thus allowing for direct tracking of transcription reactions via the fluorescent response. We suggest a set of rules that enable for the construction of responsive biomaterials that readily change their fluorescent behavior when various stimuli such as the presence of divalent ions, exposure to various nucleases, or changes in temperature are applied. We also show that the strand displacement approach can be used to program the controllable fluorescent responses in isothermal conditions. Overall, this work lays a foundation for the future development of dynamic systems for molecular computing which can be used to monitor real-time processes in cells and construct biocompatible logic gates. View Full-Text
Keywords: RNA nanotechnology; aptamer; Broccoli; dynamic nanoparticles; conditional activation RNA nanotechnology; aptamer; Broccoli; dynamic nanoparticles; conditional activation
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Chandler, M.; Lyalina, T.; Halman, J.; Rackley, L.; Lee, L.; Dang, D.; Ke, W.; Sajja, S.; Woods, S.; Acharya, S.; Baumgarten, E.; Christopher, J.; Elshalia, E.; Hrebien, G.; Kublank, K.; Saleh, S.; Stallings, B.; Tafere, M.; Striplin, C.; Afonin, K.A. Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology. Molecules 2018, 23, 3178.

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