Use of photoswitchable fluorescent proteins for droplet-based microfluidic screening

Adilya Dagkesamanskaya (1) , Krzysztof Langer (2) , Alexandra Tauzin (1) , Catherine Rouzeau (1) , Delphine Lestrade (3) , Gabrielle Potocki-Veronese (1) , Laurent Boitard (2) , Jérôme Bibette (2) , Jean Baudry (2) , Denis Pompon (1) , Véronique Le Berre (1)

1 LISBP - Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés
2 LCMD - Laboratoire Colloïdes et Matériaux Divisés
3 TWB - Toulouse White Biotechnology

11 Application of droplet-based microfluidics for the screening of microbial libraries is one of the 12 important ongoing development in functional genomics/metagenomics. In this article, we propose a 13 new method that can be employed for the high-throughput profiling of cell growth. It consists in light-14 driven labelling droplets that contain growing cells directly in microfluidics observation chamber, 15 followed by recovery of the labelled cells. This method is based on intracellular expression of green to 16 red switchable fluorescent proteins. The proof of concept is established here for two commonly used 17 biological models, E.coli and S.cerevisiae. Growth of cells in droplets was monitored under a 18 microscope and, depending on the targeted phenotype, the fluorescence of selected droplets was 19 switched from a "green" to a "red" state. Red fluorescent cells from labelled droplets were then 20 successfully detected, sorted with the Fluorescence Activated Cell Sorting machine and recovered. 21 Finally, the application of this method for different kind of screenings, in particular of metagenomic 22 libraries, is discussed and this idea is validated by the analysis of a model mini-library. 23

Mots clés: Droplet-based microfluidics; library screening; photoconvertible fluorescent protein; 24 Saccharomyces cerevisiae; Escherichia coli; Fluorescence Activated Cell Sorting 25

Domaines: Sciences du Vivant [q-bio]; Biochimie, Biologie Moléculaire

Adilya Dagkesamanskaya, Krzysztof Langer, Alexandra Tauzin, Catherine Rouzeau, Delphine Lestrade, et al.. Use of photoswitchable fluorescent proteins for droplet-based microfluidic screening. Journal of Microbiological Methods, 2018, 147, pp.59 - 65. ⟨10.1016/j.mimet.2018.03.001⟩. ⟨hal-01849551⟩