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

S. L. Sjostrom ; Y. P. Bai ; Mingtao Huang (Institutionen för kemi- och bioteknik, Systembiologi) ; Zihe Liu (Institutionen för kemi- och bioteknik, Systembiologi) ; Jens B. Nielsen (Institutionen för kemi- och bioteknik, Systembiologi) ; H. N. Joensson ; H. A. Svahn
Lab on a Chip (1473-0197). Vol. 14 (2014), 4, p. 806-813.
[Artikel, refereegranskad vetenskaplig]

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 a-amylase production, close to the theoretical maximum enrichment. Furthermore, a 105 member yeast cell library is screened yielding a clone with a more than 2-fold increase in a-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.

Nyckelord: SACCHAROMYCES-CEREVISIAE, DIRECTED EVOLUTION, MICRODROPLETS



Denna post skapades 2014-03-11. Senast ändrad 2014-10-27.
CPL Pubid: 194784

 

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Institutioner (Chalmers)

Institutionen för kemi- och bioteknik, Systembiologi (2008-2014)

Ämnesområden

Kemiteknik
Industriell bioteknik

Chalmers infrastruktur