title: Microfluidic droplet platform for ultrahigh-throughput single-cell screening of biodiversity.
abstract: Ultrahigh-throughput screening (uHTS) techniques can identify unique functionality from millions of variants. To mimic the natural selection mechanisms that occur by compartmentalization in vivo, we developed a technique based on single-cell encapsulation in droplets of a monodisperse microfluidic double water-in-oil-in-water emulsion (MDE). Biocompatible MDE enables in-droplet cultivation of different living species. The combination of droplet-generating machinery with FACS followed by next-generation sequencing and liquid chromatography-mass spectrometry analysis of the secretomes of encapsulated organisms yielded detailed genotype/phenotype descriptions. This platform was probed with uHTS for biocatalysts anchored to yeast with enrichment close to the theoretically calculated limit and cell-to-cell interactions. MDE-FACS allowed the identification of human butyrylcholinesterase mutants that undergo self-reactivation after inhibition by the organophosphorus agent paraoxon. The versatility of the platform allowed the identification of bacteria, including slow-growing oral microbiota species that suppress the growth of a common pathogen,Staphylococcus aureus, and predicted which genera were associated with inhibitory activity.
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.|Department of Chemistry, Lomonosov Moscow State University, 119991 Moscow, Russian Federation.
authors.forename: Stanislav S
authors.initials: SS
authors.lastname: Terekhov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.|Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Ivan V
authors.initials: IV
authors.lastname: Smirnov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.
authors.forename: Anastasiya V
authors.initials: AV
authors.lastname: Stepanova
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.|Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Tatyana V
authors.initials: TV
authors.lastname: Bobik
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.
authors.forename: Yuliana A
authors.initials: YA
authors.lastname: Mokrushina
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.
authors.forename: Natalia A
authors.initials: NA
authors.lastname: Ponomarenko
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation.|Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Alexey A
authors.initials: AA
authors.lastname: Belogurov
authors.affiliation: Department of Chemistry, Lomonosov Moscow State University, 119991 Moscow, Russian Federation.|Skolkovo Institute of Science and Technology, 143026 Skolkovo, Russian Federation.
authors.forename: Maria P
authors.initials: MP
authors.lastname: Rubtsova
authors.affiliation: Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Olga V
authors.initials: OV
authors.lastname: Kartseva
authors.affiliation: Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Marina O
authors.initials: MO
authors.lastname: Gomzikova
authors.affiliation: Institute of General Pathology and Pathophysiology, 125315 Moscow, Russian Federation.
authors.forename: Alexey A
authors.initials: AA
authors.lastname: Moskovtsev
authors.affiliation: St. Petersburg Academic University, 194021 St. Petersburg, Russian Federation.
authors.forename: Anton S
authors.initials: AS
authors.lastname: Bukatin
authors.affiliation: St. Petersburg Academic University, 194021 St. Petersburg, Russian Federation.
authors.forename: Michael V
authors.initials: MV
authors.lastname: Dubina
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.|Moscow Institute of Physics and Technology, 141701 Dolgoprudny, Russian Federation.
authors.forename: Elena S
authors.initials: ES
authors.lastname: Kostryukova
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Vladislav V
authors.initials: VV
authors.lastname: Babenko
authors.affiliation: Moscow Institute of Physics and Technology, 141701 Dolgoprudny, Russian Federation.
authors.forename: Maria T
authors.initials: MT
authors.lastname: Vakhitova
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Alexander I
authors.initials: AI
authors.lastname: Manolov
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Maja V
authors.initials: MV
authors.lastname: Malakhova
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Maria A
authors.initials: MA
authors.lastname: Kornienko
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.|Moscow Institute of Physics and Technology, 141701 Dolgoprudny, Russian Federation.
authors.forename: Alexander V
authors.initials: AV
authors.lastname: Tyakht
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Anna A
authors.initials: AA
authors.lastname: Vanyushkina
authors.affiliation: Research and Clinical Centre of Physical-Chemical Medicine, 119435 Moscow, Russian Federation.
authors.forename: Elena N
authors.initials: EN
authors.lastname: Ilina
authors.affiliation: Kazan Federal University, 420008 Kazan, Russian Federation.|French Academy of Pharmacy, 75270 Paris, France.
authors.forename: Patrick
authors.initials: P
authors.lastname: Masson
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russian Federation; gabibov@mx.ibch.ru sidney.altman@yale.edu.|Department of Chemistry, Lomonosov Moscow State University, 119991 Moscow, Russian Federation.|Kazan Federal University, 420008 Kazan, Russian Federation.
authors.forename: Alexander G
authors.initials: AG
authors.lastname: Gabibov
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06520 gabibov@mx.ibch.ru sidney.altman@yale.edu.
authors.forename: Sidney
authors.initials: S
authors.lastname: Altman
mesh_terms: D000898:Antibiosis; D044822:Biodiversity; D002091:Butyrylcholinesterase / Q000737:chemistry*; D002450:Cell Communication; D004655:Emulsions; D005434:Flow Cytometry; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing; D057166:High-Throughput Screening Assays / Q000295:instrumentation*; D006801:Humans; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D009822:Oils, Volatile / Q000737:chemistry; D010261:Paraoxon / Q000737:chemistry*; D010641:Phenotype; D059010:Single-Cell Analysis / Q000295:instrumentation*; D013211:Staphylococcus aureus / Q000187:drug effects / Q000254:growth & development; D014867:Water / Q000737:chemistry
chemical_list: D004655:Emulsions; D009822:Oils, Volatile; D014867:Water; D002091:Butyrylcholinesterase; D010261:Paraoxon
keywords: Staphylococcus aureus; butyrylcholinesterase; cell–cell interactions; microfluidic encapsulation; ultrahigh-throughput screening