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To improve β-ketovaleryl-CoA pools for further increase of OCFA production, we co-expressed the bktB encoding β-ketothiolase in the producing strain, and the OCFA production was increased by 33% compared to control. Combining strain engineering and the optimization of the C/N ratio promoted the OCFA production up to 1.87 g/L representing 62% of total lipids, the highest recombinant OCFAs titer reported in yeast, up to date. This study provides a strong basis for the microbial production of OCFAs and its derivatives having high potential