1. Optimization of fluorogenic RNA-based biosensors using droplet-based microfluidic ultrahigh-throughput screening
- Author
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Roger Cubi, Farah Bouhedda, Michael Ryckelynck, Alexis Autour, Architecture et Réactivité de l'ARN (ARN), Institut de biologie moléculaire et cellulaire (IBMC), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Plate-Forme de Recherche en Imagerie Cellulaire de Haute-Normandie (PRIMACEN), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Institute for Research and Innovation in Biomedicine (IRIB), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), WERLING, Danièle, Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-High-tech Research Infrastructures for Life Sciences (HeRacLeS), and Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
light-up aptamer ,[SDV.BIO]Life Sciences [q-bio]/Biotechnology ,Computer science ,High-throughput screening ,Microfluidics ,fluorogenic biosensors ,Biosensing Techniques ,Signal ,high-throughput screening ,General Biochemistry, Genetics and Molecular Biology ,Domain (software engineering) ,03 medical and health sciences ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,Molecular Biology ,Throughput (business) ,[SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,Fluorescent Dyes ,030304 developmental biology ,chemistry.chemical_classification ,next generation sequencing ,0303 health sciences ,Event (computing) ,Biomolecule ,030302 biochemistry & molecular biology ,High-Throughput Nucleotide Sequencing ,aptasensors ,[SDV.BIO] Life Sciences [q-bio]/Biotechnology ,chemistry ,RNA ,Biological system ,Biosensor - Abstract
International audience; 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.
- Published
- 2019