Ultra-high-throughput picoliter-droplet microfluidics screening of the industrial cellulase-producing filamentous fungus Trichoderma reesei

Journal of Industrial Microbiology & Biotechnology
Ronglin HeRan Tu

Abstract

The selection of improved producers among the huge number of variants in mutant libraries is a key issue in filamentous fungi of industrial biotechnology. Here, we developed a droplet-based microfluidic high-throughput screening platform for selection of high-cellulase producers from filamentous fungus Trichoderma reesei. The screening system used a fluorogenic assay to measure amount of cellulase and its activity. The key effectors such as cellulase-inducing medium, spore germination, droplet cultivation time, droplet fluorescence signal detection, and droplet cell sorting were studied. An artificial pre-mixed library of high- and low-cellulase-producing T. reesei strains was screened successfully to verify the feasibility of our method. Finally, two cellulase hyperproducers exhibiting improvements in cellulase activity of 27% and 46% were isolated from an atmospheric and room-temperature plasma (ARTP)-mutated library. This high-throughput screening system could be applied to the engineering of T. reesei strains and other industrially valuable protein-producing filamentous fungi.

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Citations

Apr 24, 2020·Frontiers in Bioengineering and Biotechnology·Qin WangHan Xiao
Sep 28, 2020·Journal of Industrial Microbiology & Biotechnology·Ran TuQinhong Wang
Dec 2, 2020·International Journal of Biological Macromolecules·Patricia P AdrianiFelipe S Chambergo
Mar 19, 2021·Critical Reviews in Biotechnology·Jianhua YangLeilei Zhu
Nov 28, 2021·Scientific Reports·Sehrish IftikharJulia Elisa Sepulveda-Diaz

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Methods Mentioned

BETA
FACS
fluorescence-activated dielectrophoresis
PCR
phosphotransferase
fluorescence microscopy

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