Crispr-based multi-gene integration strategies to create saccharomyces cerevisiae strains for consolidated bioprocessing

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dc.contributor.authorJacob, Odwa
dc.contributor.authorvan Lill, Gert Rutger
dc.contributor.authorden Haan, Riaan
dc.date.accessioned2023-02-10T08:08:22Z
dc.date.available2023-02-10T08:08:22Z
dc.date.issued2022
dc.description.abstractSignificant engineering of Saccharomyces cerevisiae is required to enable consolidated bioprocessing (CBP) of lignocellulose to ethanol. Genome modification in S. cerevisiae has been successful partly due to its efficient homology-directed DNA repair machinery, and CRISPR technology has made multi-gene editing even more accessible. Here, we tested the integration of cellulase encoding genes to various sites on the yeast genome to inform the best strategy for creating cellulolytic strains for CBP. We targeted endoglucanase (EG) or cellobiohydrolase (CBH) encoding genes to discreet chromosomal sites for single-copy integration or to the repeated delta sites for multi-copy integration. CBH1 activity was significantly higher when the gene was targeted to the delta sequences compared to single gene integration loci. EG production was comparable, though lower when the gene was targeted to a chromosome 10 site. We subsequently used the information to construct a strain containing three cellulase encoding genes. While individual cellulase activities could be assayed and cellulose conversion demonstrated, it was shown that targeting specific genes to specific loci had dramatic effects on strain efficiency. Since marker-containing plasmids could be cured from these strains, additional genetic changes can subsequently be made to optimize strains for CBP conversion of lignocellulose.en_US
dc.identifier.citationJacob, O. et al. (2022). Crispr-based multi-gene integration strategies to create saccharomyces cerevisiae strains for consolidated bioprocessing. Applied Sciences, 12(23), 12317. https://doi.org/10.3390/app122312317en_US
dc.identifier.issn2076-3417
dc.identifier.urihttps://doi.org/10.3390/app122312317
dc.identifier.urihttp://hdl.handle.net/10566/8406
dc.language.isoenen_US
dc.publisherMDPIen_US
dc.subjectBiotechnologyen_US
dc.subjectSaccharomyces cerevisiaeen_US
dc.subjectYeasten_US
dc.subjectFood productionen_US
dc.titleCrispr-based multi-gene integration strategies to create saccharomyces cerevisiae strains for consolidated bioprocessingen_US
dc.typeArticleen_US

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