Ultrahigh-throughput screening of Trichoderma reesei strains capable of carbon catabolite repression release and cellulase hyperproduction using a microfluidic droplet platform.

Xuan Chinh Luu, Yosuke Shida, Yoshiyuki Suzuki, Daiki Kuwahara, Takeshi Fujimoto, Yuka Takahashi, Naomi Sato, Akihiro Nakamura, Wataru Ogasawara

Bioscience, biotechnology, and biochemistry 2023 Oct 25

filter terms:

Trichoderma reesei is the most well-known cellulase producer in the biorefinery industry. Its cellulase biosynthesis is repressed by glucose via carbon catabolite repression (CCR), making CCR-releasing strains with cellulase hyperproduction desirable. Here, we employed a microfluidic droplet platform to culture and screen T. reesei mutants capable of CCR release and cellulase overproduction from extensive mutagenesis libraries. With 3 mutagenesis rounds, about 6.20 × 103 droplets were sorted from a population of 1.51 × 106 droplets in a period of 4.4 h; 76 recovery mutants were screened on flask fermentation, and 2 glucose uptake retarded mutants, MG-9-3 and MG-9-3-30, were eventually isolated. We also generated a hypercellulase producer, M-5, with CCR release via a single mutagenesis round. The hyphal morphology and molecular mechanisms in the mutants were analyzed. This versatile approach combined with a comprehensive understanding of CCR release mechanisms will provide innovative and effective strategies for low-cost cellulase production. © The Author(s) 2023. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.

Citation

Xuan Chinh Luu, Yosuke Shida, Yoshiyuki Suzuki, Daiki Kuwahara, Takeshi Fujimoto, Yuka Takahashi, Naomi Sato, Akihiro Nakamura, Wataru Ogasawara. Ultrahigh-throughput screening of Trichoderma reesei strains capable of carbon catabolite repression release and cellulase hyperproduction using a microfluidic droplet platform. Bioscience, biotechnology, and biochemistry. 2023 Oct 25;87(11):1393-1406