A barrier to cost-efficient biomanufacturing is the instability of engineered genetic elements, such as plasmids. Instability can also manifest at the whole-genome level, when fungal dikaryons revert to parental species due to nuclear segregation during cell division. Here, we show that by encapsulating Saccharomyces cerevisiae-Pichia stipitis dikaryons in an alginate matrix, we can limit cell division and preserve their expanded metabolic capabilities. As a proxy to cellulosic ethanol production, we tested the capacity of such cells to carry out ethanologenic fermentation of glucose and xylose, examining substrate use, ploidy, and cell viability in relation to planktonic fusants, as well as in relation to planktonic and encapsulated cell cultures consisting of mixtures of these species. Glucose and xylose consumption and ethanol production by encapsulated dikaryons were greater than planktonic controls. Simultaneous co-fermentation did not occur; rather the order and kinetics of glucose and xylose catabolism by encapsulated dikaryons were similar to cultures where the two species were encapsulated together. Over repeated cycles of fed-batch culture, encapsulated S. cerevisiae-P. stipitis fusants exhibited a dramatic increase in genomic stability, relative to planktonic fusants. Encapsulation also increased the stability of antibiotic-resistance plasmids used to mark each species and preserved a fixed ratio of S. cerevisiae to P. stipitis cells in mixed cultures. Our data demonstrate how encapsulating cells in an extracellular matrix restricts cell division and, thereby, preserves the stability and biological activity of entities ranging from genomes to plasmids to mixed populations, each of which can be essential to cost-efficient biomanufacturing.

中文翻译：

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封装增强了原生质体融合体的稳定性。

具有成本效益的生物制造的一个障碍是工程遗传元件（如质粒）的不稳定性。不稳定性也可以在全基因组水平上表现出来，当真菌双核由于细胞分裂过程中的核分离而恢复到亲本物种时。在这里，我们表明通过将酿酒酵母-树干毕赤酵母双核封装在藻酸盐基质中，我们可以限制细胞分裂并保持其扩展的代谢能力。作为纤维素乙醇生产的代表，我们测试了这些细胞进行葡萄糖和木糖乙醇发酵的能力，检查底物使用、倍性和与浮游融合体相关的细胞活力，以及与浮游和封装细胞相关的细胞活力由这些物种的混合物组成的培养物。封装双核的葡萄糖和木糖消耗量和乙醇产量高于浮游对照。未发生同时共发酵；相反，封装双核的葡萄糖和木糖分解代谢的顺序和动力学类似于将这两种物质封装在一起的培养物。经过重复的补料分批培养循环，封装的酿酒酵母-P。相对于浮游融合子，树干融合子表现出基因组稳定性的显着增加。封装还增加了用于标记每个物种的抗生素抗性质粒的稳定性，并在混合培养物中保持了酿酒酵母与树干假单胞菌细胞的固定比例。我们的数据证明了将细胞封装在细胞外基质中如何限制细胞分裂，从而，