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Lab Chip. 2014 Feb 21;14(4):806-13. doi: 10.1039/c3lc51202a.

High-throughput screening for industrial enzyme production hosts by droplet microfluidics.

Author information

1
Division of Proteomics and Nanobiotechnology, Science for Life Laboratory, Royal Institute of Technology (KTH), Sweden. helenea@biotech.kth.se.

Abstract

A high-throughput method for single cell screening by microfluidic droplet sorting is applied to a whole-genome mutated yeast cell library yielding improved production hosts of secreted industrial enzymes. The sorting method is validated by enriching a yeast strain 14 times based on its α-amylase production, close to the theoretical maximum enrichment. Furthermore, a 10(5) member yeast cell library is screened yielding a clone with a more than 2-fold increase in α-amylase production. The increase in enzyme production results from an improvement of the cellular functions of the production host in contrast to previous droplet-based directed evolution that has focused on improving enzyme protein structure. In the workflow presented, enzyme producing single cells are encapsulated in 20 pL droplets with a fluorogenic reporter substrate. The coupling of a desired phenotype (secreted enzyme concentration) with the genotype (contained in the cell) inside a droplet enables selection of single cells with improved enzyme production capacity by droplet sorting. The platform has a throughput over 300 times higher than that of the current industry standard, an automated microtiter plate screening system. At the same time, reagent consumption for a screening experiment is decreased a million fold, greatly reducing the costs of evolutionary engineering of production strains.

PMID:
24366236
DOI:
10.1039/c3lc51202a
[Indexed for MEDLINE]
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