Patent FR-2959514-A1 (FR)
Title: Preparing an industrial yeast strain Saccharomyces cerevisiae, comprises selecting strain, integrating an expression cassette/deletion of a cassette and inducing expression of a gene and deleting at least two copies of open reading frame
Inventor: DESFOUGERES THOMAS
IPC: C12N1/19
Filed: 2010-04-30
Abstract: Preparing an industrial yeast strain Saccharomyces cerevisiae, comprises: (i) selecting and obtaining the strain; (ii) integrating an expression cassette or deletion, in the genome of the yeast, of a cassette e.g. (a) combination of open reading frame (ORF) of the gene of Pichia stipitisXRm encoding for the mutated reductase xylose enzyme; (iii) inducing the expression of at least one gene from each step of the non-oxidative part of pentose phosphate pathway; and (iv) deleting at least two copies of ORF of Saccharomyces cerevisiaeGRE3 gene. Process for preparing an industrial yeast strain Saccharomyces cerevisiaeto produce ethanol from a medium comprising at least one pentose, comprises: (i) selecting and obtaining an industrial yeast strain of Saccharomyces cerevisiaecapable of producing high concentrations of ethanol, at least 14.5% (v/v), preferably at least 16% (v/v) on hydrolyzate of cereals, in conditions of simultaneous saccharification and fermentation (SSF) and at a temperature of 35[deg] C; (ii) integrating at least one expression cassette or deletion, in the genome of the yeast of step (i), at least one cassette such as (a) the combination of open reading frame (ORF) of the gene of Pichia stipitisXRm encoding for the mutated reductase xylose enzyme using the NADH, H+ as a cofactor preferably in place and instead of NADPH, H+/promoter and terminator of Saccharomyces cerevisiae, where the cassette is flanked upstream and downstream of recombinogenic regions allowing its integration targeted in the genome, (b) the combination of open reading frame (ORF) of Pichia stipitisXDH gene coding for the xylitol dehydrogenase enzyme/promoter and terminator of Saccharomyces cerevisiae, where the cassette is flanked upstream and downstream of recombinogenic regions allowing its targeted integration into the genome, (c) combination of open reading frame (ORF) of the gene of Saccharomyces cerevisiaeXKS1 encoding the xylulokinase enzyme/promoter and terminator of Saccharomyces cerevisiae, where the cassette is flanked upstream and downstream of recombinogenic regions allowing its integration targeted into the genome; (iii) inducing the expression of at least one gene from each step of the non-oxidative part of pentose phosphate pathway by placing it under the control of a promoter of a gene of the glycolysis strongly expressed during alcoholic fermentation; and (iv) deleting at least two copies of the open reading frame (ORF) of Saccharomyces cerevisiaeGRE3 gene encoding aldose dehydrogenase. Independent claims are included for: (1) yeast strain Saccharomyces cerevisiaeEG3 deposited under the NO: l-4295 at the CNCM; (2) yeast strain Saccharomyces cerevisiaeEG2 deposited under the NO: l-4294 at the CNCM and the strain resulting from the evolution directed strain EG3; (3) yeast strain Saccharomyces cerevisiaeEG1 deposited under the NO: l-4293 at the CNCM, and strain resulting from the directed evolution of the strain EG3; (4) yeast strain Saccharomyces cerevisiaeEG9 deposited under the NO: l-4450 at the CNCM; and #a process for producing ethanol, from a medium comprising at least one pentose, comprising fermenting the yeast.