Peptone

The yeast strain GRX1-GFP-HIS3MX6 was purchased from Thermo Ficher Scientific. All the primers used in this study are listed in Supplementary Table 1. To create the GRX1-GFP-tdTomato-kanR strain, a PCR fragment containing tdTomato-kanR and homologies to GFP-HIS3MX6 was amplified with primers C1 and C2 from pfa6a-tdTomato-kanR (constructed in our lab) and transformed to the GRX1-GFP-HIS3MX6 strain.
All the strains were grown in liquid YPD medium containing 20 g/L glucose (Sigma), 10 g/L peptone (Euromedex) and 10 g/L yeast extraction (Euromedex). When needed, 20 g/L agar (Euromedex) was added to make solid plates. YNB-URA⁻ plates [20 g/L glucose (Sigma), 20 g/L agar (Euromedex), 1.71 g/L yeast nitrogen base without amino acids and nitrogen (Euromedex), 5 g/L ammonium sulfate (Sigma), and 0.77 g/L CSM-URA⁻ (Euromedex)] or 5-FOA plate [20 g/L glucose (Sigma), 20 g/L agar (Euromedex), 1.71 g/L yeast nitrogen base without amino acids and nitrogen (Euromedex), 5 g/L ammonium sulfate (Sigma), 0.79 g/L CSM-URA⁻ (Euromedex), and 1 g/L 5-FOA (Euromedex)] were used to select transformants.

The Y. lipolytica quadruple mutant strain JMY1877 (MATA leu2-270 ura3-302 Δdga1 Δlro1 Δare1 Δdga2) [5 (link)] was transformed using the lithium acetate method [33 ] either with the pTEF-EgDGAT1-1-URA3, the pTEF-AtDGAT1-URA3 cassette or the empty pTEF-URA3 cassette (control strain) from the NotI linearized JMP62 recombinant and native vectors. Yeast were grown in uracil deficient medium containing 0.17% (w/v) yeast nitrogen base (YNB) supplemented with 0.5% (w/v) ammonium sulfate and 0.2% (w/v) casamino acids (BD, Le Pont de Claix, France) and YP medium containing 2.2% (w/v) peptone and 1.1% (w/v) yeast extract (Euromedex, Mundolsheim, France). The carbon sources (Sigma-Aldrich, Saint-Quentin Fallavier, France) were 2% (w/v) glucose for YPG and YNBG media or 0.02% (w/v) glucose and 2% (w/v) lauric acid methyl ester (LAME) emulsified by sonication with 0.2% (w/v) Tween 20 for YPL medium. LAME was used instead of lauric acid which is solid at temperatures below 44°C. Transformants were selected on YNBG plates. Cells from three independent transformants were grown for one day in YPG medium. Yeasts were diluted in YPG or YPL medium to an optical density of 0.5 for an 18h culture. All cultures were performed in baffled Erlenmeyer flasks at 28°C and 200 r.p.m.