Data_Sheet_2_Lager Yeast Design Through Meiotic Segregation of a Saccharomyces cerevisiae × Saccharomyces eubayanus Hybrid.ZIP

Yeasts in the lager brewing group are closely related and consequently do not exhibit significant genetic variability. Here, an artificial Saccharomyces cerevisiae × Saccharomyces eubayanus tetraploid interspecies hybrid was created by rare mating, and its ability to sporulate and produce viable gametes was exploited to generate phenotypic diversity. Four spore clones obtained from a single ascus were isolated, and their brewing-relevant phenotypes were assessed. These F1 spore clones were found to differ with respect to fermentation performance under lager brewing conditions (15°C, 15 °Plato), production of volatile aroma compounds, flocculation potential and temperature tolerance. One spore clone, selected for its rapid fermentation and acetate ester production was sporulated to produce an F2 generation, again comprised of four spore clones from a single ascus. Again, phenotypic diversity was introduced. In two of these F2 clones, the fermentation performance was maintained and acetate ester production was improved relative to the F1 parent and the original hybrid strain. Strains also performed well in comparison to a commercial lager yeast strain. Spore clones varied in ploidy and chromosome copy numbers, and faster wort fermentation was observed in strains with a higher ploidy. An F2 spore clone was also subjected to 10 consecutive wort fermentations, and single cells were isolated from the resulting yeast slurry. These isolates also exhibited variable fermentation performance and chromosome copy numbers, highlighting the instability of polyploid interspecific hybrids. These results demonstrate the value of this natural approach to increase the phenotypic diversity of lager brewing yeast strains.

拉格啤酒酿造菌群中的酵母亲缘关系紧密，因此未表现出显著的遗传变异。本研究通过罕见交配构建了人工酿酒酵母（Saccharomyces cerevisiae）×真贝酵母（Saccharomyces eubayanus）四倍体种间杂种，并利用其产孢及产生有活力配子的能力来生成表型多样性。从单个子囊中分离得到4个孢子克隆，并评估了其与啤酒酿造相关的表型。研究发现，这些F₁代孢子克隆在拉格酿造条件（15℃、15° Plato）下的发酵性能、挥发性香气物质生成能力、絮凝潜能以及温度耐受性方面存在差异。选取其中一株发酵速度快且乙酸酯产量高的孢子克隆进行产孢，获得F₂代群体，同样从单个子囊中分离得到4个孢子克隆，再次引入了表型多样性。其中两株F₂代克隆保留了亲本的发酵性能，且乙酸酯产量相较于F₁代亲本及原始杂种菌株得到提升，其表现亦优于商业拉格酵母菌株。孢子克隆的倍性及染色体拷贝数存在差异，且倍性更高的菌株麦汁发酵速度更快。选取一株F₂代孢子克隆进行10轮连续麦汁发酵，并从后续的酵母泥中分离单细胞。这些分离株同样表现出不同的发酵性能与染色体拷贝数，凸显了多倍体种间杂种的基因组不稳定性。本研究结果证实了该自然策略在提升拉格啤酒酿造酵母菌株表型多样性方面的应用价值。