Proc Natl Acad Sci U S A, 2017, 114(10):2550-2555

Microfluidic droplet platform for ultrahigh-throughput single-cell screening of biodiversity

All rights reserved. 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.

Terekhov SS, Smirnov IV, Stepanova AV, Bobik TV, Mokrushina YA, Ponomarenko NA, Belogurov AA, Rubtsova MP, Kartseva OV, Gomzikova MO, Moskovtsev AA, Bukatin AS, Dubina MV, Kostryukova ES, Babenko VV, Vakhitova MT, Manolov AI, Malakhova MV, Kornienko MA, Tyakht AV, Vanyushkina AA, Ilina EN, Masson P, Gabibov AG, Altman S

IBCH: 3300
Ссылка на статью в журнале: http://www.pnas.org/lookup/doi/10.1073/pnas.1621226114
Кол-во цитирований на 07.2019: 33
Данные статьи проверены модераторами 2017-03-07