CRISPR–Cas system enables fast and simple genome editing of industrial Saccharomyces cerevisiae strains

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Stovicek, Vratislav ; Borodina, Irina ; Förster, Jochen (2015)
  • Publisher: Elsevier BV
  • Journal: volume 2, pages 13-22 (issn: 2214-0301)
  • Related identifiers: doi: 10.1016/j.meteno.2015.03.001
  • Subject: Genome editing | Biotechnology | Chemical production | Industrial yeast | CRISPR–Cas9 | Biorefineries | Biology (General) | TP248.13-248.65 | QH301-705.5

There is a demand to develop 3rd generation biorefineries that integrate energy production with the production of higher value chemicals from renewable feedstocks. Here, robust and stress-tolerant industrial strains of Saccharomyces cerevisiae will be suitable production organisms. However, their genetic manipulation is challenging, as they are usually diploid or polyploid. Therefore, there is a need to develop more efficient genetic engineering tools. We applied a CRISPR–Cas9 system for genome editing of different industrial strains, and show simultaneous disruption of two alleles of a gene in several unrelated strains with the efficiency ranging between 65% and 78%. We also achieved simultaneous disruption and knock-in of a reporter gene, and demonstrate the applicability of the method by designing lactic acid-producing strains in a single transformation event, where insertion of a heterologous gene and disruption of two endogenous genes occurred simultaneously. Our study provides a foundation for efficient engineering of industrial yeast cell factories. Keywords: CRISPR–Cas9, Genome editing, Industrial yeast, Biorefineries, Chemical production
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