Digitalization drives accelerated process optimization by comprehensive automation. In the advanced biofuels sector this demands automatable high-throughput processes for production strain development and downstream process performance monitoring. In that context, the unit operations of oleaginous yeast-based biodiesel production are amenable to high-throughput process development. Cutaneotrichosporon oleaginosus is a leading production strain for high-energy biofuel options, that is capable of utilizing a broad range of substrates as carbon sources, thereby generating in excess of 60% (w/w) lipids under nutrient limiting conditions. For the first time, we report on the use of fast neutron (FN) irradiation for the rapid, high-throughput genetic enhancement of an oleaginous yeast in conjunction with high-throughput selection of enhanced lipid producing C. oleaginosus mutants by cultivation in the presence of the fatty acid biosynthesis inhibitor cerulenin. Performance monitoring of improved mutants was accomplished by development of a high-throughput lipid qualification methodology based on a miniaturized, low cost Nile red based spectrofluorimetric assay. From the FN mutant library, this high-throughput strain development approach allowed identification of a C. oleaginosus variant (FN M2) displaying a 21.67% (w/v) and 22.58% (w/v) increase in biomass formation and total lipid yield compared to wild-type strain, respectively. Mutant triglyceride characterization revealed a higher content of saturated fatty acids, which is favorable with respect to biofuels production standards, determined here for the first time. This study is an initial step towards an automatable, high-throughput yeast oil optimization process that facilitates accelerated industrial deployment.

中文翻译：

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朝着微生物脂质生产的高通量优化：从菌株开发到过程监控

数字化通过全面的自动化来推动流程优化。在先进的生物燃料领域，这需要用于生产菌株开发和下游过程性能监控的自动化高通量过程。在这种情况下，基于油质酵母的生物柴油生产的单元操作适合高通量工艺开发。猪角皮猪孢是高能生物燃料选择的领先生产菌株，能够利用多种底物作为碳源，从而在营养限制条件下产生超过60％（w / w）的脂质。首次，我们报道了使用快速中子（FN）辐射对油性酵母进行快速，高通量遗传增强，并结合了高通量选择产生脂质的产油梭菌。通过在脂肪酸生物合成抑制剂铜绿素的存在下培养获得突变体。通过开发基于微型低成本低成本尼罗红的分光荧光测定法的高通量脂质鉴定方法，可以完成对改良突变体的性能监控。从FN突变体文库中，这种高通量菌株开发方法可鉴定油酸梭菌。变体（FN M2）与野生型菌株相比，分别显示出21.68％（w / v）和22.58％（w / v）的生物量形成和总脂质产量增加。突变型甘油三酸酯的表征显示出较高的饱和脂肪酸含量，这相对于此处首次确定的生物燃料生产标准而言是有利的。这项研究是朝着自动化，高通量酵母油优化过程迈出的第一步，该过程有助于加速工业部署。