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Improving CRISPR-Cas9 mediated genome integration in interspecific hybrid yeasts. The uploaded files contains the experimental data and their processingof the figures used in the manuscript.
Saccharomyces pastorianus is not a classical taxon, it is an interspecific hybrid resulting from the cross of Saccharomyces cerevisiae and Saccharomyces eubayanus. Exhibiting heterosis for phenotypic traits as wort a-oligosacharides consumption and fermentation at low temperature, it has been domesticated to become the main work-horse of the brewing industry.
Although CRISPR-Cas9 has been shown to be functional in S. pastorianus, repair of CRISPR induced double strand break is unpredictable and preferentially uses the homoeologous chromosome as template preventing targeted introduction of the desired repair construct. Here, we demonstrate that lager hybrids can be edited with near 100% efficiency at carefully selected landing sites on the chimeric SeScCHRIII. The landing sites were systematically selected and evaluated for i) absence of loss of heterozygosity upon CRISPR-editing, ii) efficiency of the gRNA, and iii) absence of effect on strain physiology. Successful examples of highly efficient single and double genes integration illustrated that genome editing can be applied in interspecies hybrids paving the way to a new impulse to lager yeast strain development.
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