Development of mutant microalgae that accumulate lipids under nitrate-replete conditions

Microalgal biofuels have attracted global attention as an alternative energy source to fossil fuels. Microalgae generally accumulate triacylglycerols (lipids) under nitrogen-deficient conditions; however, cell growth is highly suppressed under these conditions, subsequently reducing lipid productivity. Therefore, microalgae that can accumulate lipids even under nitrogen-replete conditions are needed to realize the one-step nutrient-replete cultivation with improved lipid productivity. Thus, the aim of this study was to develop a method for screening mutant microalgae showing high lipid accumulation in the presence of a nitrogen source. Mutant cells were generated by irradiating the oleaginous green microalga Chlamydomonas sp. KOR1 with carbon-ion beams, followed by cultivation under nitrate-replete conditions, and then subjected to fluorescence-activated cell sorting-based screening for detecting lipid-rich cells. By repeatedly performing the sequential procedures of cultivation and selection, the strains KAC1710 and KAC1801, which showed high lipid accumulation under nitrate-replete conditions, were obtained. In the presence of abundant nitrate, these mutants formed a large number of lipid droplets in the cells, and achieved 1.5-fold (KAC1710) and 2.1-fold (KAC1801) higher lipid content than KOR1. Metabolome analysis revealed that the pool size of pyruvate was significantly increased in KAC1710 and KAC1801, consistent with the activation of lipid synthesis under nitrate-replete conditions. In conclusion, this study reports the use of a novel nitrogen-based screening method for developing mutant microalgae that accumulate lipids in the presence of a nitrogen source, and found that an increase in pyruvate pool size is an important metabolic phenomenon associated with lipid production. •Mutant microalgae that accumulate lipids under nitrate-replete conditions are needed.•Chlamydomonas sp. mutant KAC1801 can accumulate lipids in nitrogen-replete conditions.•KAC1801 accumulated 2.1-fold higher lipid than the parent strain.•Pyruvate accumulation observed in KAC1801 under nitrate-replete conditions.