Under nutrient-limited conditions, the red yeast Rhodosporidium toruloides can accumulate neutral lipids, of which the compositional fatty acids are mainly saturated and mono-unsaturated ones with 16 or 18 carbon atoms. To improve the linoleic acid content in the lipids, we enabled galactose-inducible expression of the gene encoding Delta 12-fatty acid desaturase (FADS) from Mortierella alpina or Fusarium verticillioides by integration of the corresponding expression cassettes into the genome of R. toruloides haploid and diploid strains. The relative linoleic acid content increased up to fivefold and the final linoleic acid titer reached 1.3 g/L under flask culture conditions. Our results suggested that R. toruloides may be further explored as cell factory for production of high-valued lipids and other fatty acid derivatives as bio-based chemicals and fuels.