Fig. 3: ¹³C-methanol fermentations to characterise the reconstructed evolved strain.

a Growth profile of CEN.PK 113-5D and the reconstructed evolved strain grown in liquid YNB medium with 2% ¹³C-methanol supplemented with 0.1% yeast extract cultures in bioreactors. Data are from two independent biological replicates with lines representing the mean. Source data are provided as a Source Data file. b Percentage of ¹³C-CO₂/CO₂ production in yeast extract ¹³C-methanol (2%) medium from two biological replicates with bars representing the mean. c ¹³C-ethanol was produced by CEN.PK 113-5D and by the reconstructed evolved strain. The signal intensity was normalised to the inert gas nitrogen, and then to biomass for each strain. Data show ¹³C-ethanol intensity at 47 amu for two independent biological replicates, with bars representing the mean. d Comparison of ¹²C-ethanol and ¹³C-ethanol intensities during growth in ¹³C-methanol. The signal intensity was normalised to the inert gas nitrogen, and then to biomass for each strain. Data show the ¹²C-ethanol and ¹³C-ethanol intensity at 31 and 33 amu, respectively, for two independent biological replicates, with bars representing means. SEM secondary electron multiplier, amu atomic mass unit. e Percentage of fully ¹³C-labelled intracellular metabolite pool relative to total metabolite pool of CEN.PK 113-5D and reconstructed evolved strains. F1,6BP fructose-1,6-bisphosphate, G3P glyceraldehyde-3-phosphate, RL5P ribulose-5-phosphate, DHAP dihydroxyacetone phosphate, CIT citrate, 3PG 3-phosphoglyceric acid, PEP phosphoenolpyruvate, ACCoA acetyl-coenzyme A. Data are from two independent biological replicates, with bars representing means, except for CEN.PK 113-5D F1,6BP, for which no 12 C metabolite was detected in one replicate. Purple squares represent the Reconstructed EC strain, and black circles represent the CEN.PK 113-5D strain. Source data are provided as a Source Data file.