Yarrowia lipolytica recombinant strain GGY237 bearing a heterologous gene encoding alpha-amylase SoA (S itophilus o ryzae alpha-a mylase; GenBank: KP027641.1) was used in this study [26] (link). This Po1h-derived strain (MatA, ura3–302, xpr2–322, axp1–2) was constructed using Golden Gate modular cloning strategy, as described previously [27] (link). SoA gene was transcriptionally fused to a signal peptide of YALI0B03564g gene for efficient secretion of the protein [28] (link) and flanked with 4UASpTEF promoter and tLip2 terminator.
Precultures of Y. lipolytica GGY237 were developed from 15% glycerol stocks stored at −80 °C, prior to each culture run. The biomass was spread on YPD agar medium containing (g L−1): yeast extract (YE; Merck, USA), 10; bactopeptone (BP; Merck, USA), 20; glucose (POCH, Poland), 20; agar (BIOCORP, Poland), 20. Subsequently, 24‐h biomass was transferred into 1‐L Erlenmeyer flasks containing 100 ml of YPG20 medium and incubated at 30 °C with 250 rpm agitation in a rotary shaker (BIOSAN, Latvia) for 23 h.
Precultures of Y. lipolytica GGY237 were developed from 15% glycerol stocks stored at −80 °C, prior to each culture run. The biomass was spread on YPD agar medium containing (g L−1): yeast extract (YE; Merck, USA), 10; bactopeptone (BP; Merck, USA), 20; glucose (POCH, Poland), 20; agar (BIOCORP, Poland), 20. Subsequently, 24‐h biomass was transferred into 1‐L Erlenmeyer flasks containing 100 ml of YPG20 medium and incubated at 30 °C with 250 rpm agitation in a rotary shaker (BIOSAN, Latvia) for 23 h.