Journal
METHODS
Volume 161, Issue -, Pages 46-53Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymeth.2019.03.015
Keywords
Fluorogenic biosensors; Light-up aptamer; High-throughput screening; Next generation sequencing; RNA; Aptasensors
Funding
- Agence Nationale de la Recherche (BrightRiboProbes) [ANR-16-CE11-0010-01/02]
- University of Strasbourg Institute of Advanced Study (USIAS, program Translatomix)
- LabEx [ANR-10-LABX-0036_NETRNA]
- Universite de Strasbourg
- Centre National de la Recherche Scientifique
- French National Research Agency
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Biosensors are biological molecules able to detect and report the presence of a target molecule by the emission of a signal. Nucleic acids are particularly appealing for the design of such molecule since their great structural plasticity makes them able to specifically interact with a wide range of ligands and their structure can rearrange upon recognition to trigger a reporting event. A biosensor is typically made of three main domains: a sensing domain that is connected to a reporting domain via a communication module in charge of transmitting the sensing event through the molecule. The communication module is therefore an instrumental element of the sensor. This module is usually empirically developed through a trial-and-error strategy with the testing of only a few combinations judged relevant by the experimenter. In this work, we introduce a novel method combining the use of droplet-based microfluidics and next generation sequencing. This method allows to functionally characterize up to a million of different sequences in a single set of experiments and, by doing so, to exhaustively test every possible sequence permutations of the communication module. Here, we demonstrate the efficiency of the approach by isolating a set of optimized RNA biosensors able to sense theophylline and to convert this recognition into fluorescence emission.
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