Adaptive Evolution of Rhodosporidium toruloides with Ultra-centrifugal Fractionation Leads to a High Microbial Lipid...

In situ reintegration of crude glycerol into lipid feedstock can increase the production efficiency and environmental tolerance for the biodiesel industry. In this study, a wild oleaginous yeast Rhodosporidium toruloides CGMCC 2.1389 was selected for its high cell growth and lipid accumulation in crude glycerol. Ultra-centrifugal fractionation was applied in a long-term adaptive evolution, and a stable R. toruloides XR20 strain was finally obtained. The lipid content and titer of the R. toruloides XR20 strain in flask increased by 100.1% and 75.8% compared with the parental strain. Record-breaking lipid titers using crude glycerol by R. toruloides XR20 were achieved by batch and fed-batch fermentation, reaching 24.5 ± 0.5 g/L and 42.5 ± 0.8 g/L, respectively. R. toruloides XR20 showed enlarged intracellular space, higher intracellular NADPH, and acetyl-CoA contents compared with the parental strain. The changes in the expression levels of the key genes favored lipid synthesis in R. toruloides XR20. This study constructed a robust high lipid-producing R. toruloides on crude glycerol, contributing to a glycerol-free biodiesel production process.