Additional file 1 of Enhanced upgrading of lignocellulosic substrates by coculture of Saccharomyces cerevisiae and Acinetobacter baylyi ADP1
Creators: name: Liu, Changshuo
name: Choi, Bohyun
name: Efimova, Elena
name: Nygård, Yvonne
name: Santala, Suvi
Publisher: name: figshare
Year: 2024
Subjects: subject: Biochemistry
subject: Space Science
subject: Medicine
subject: Microbiology
subject: FOS: Biological sciences
subject: Genetics
subject: Molecular Biology
subject: Pharmacology
subject: Biotechnology
subject: Chemical Sciences not elsewhere classified
subject: Ecology
subject: Biological Sciences not elsewhere classified

description: Additional file 1: Table S1. Primers used in this study. Table S2. Plasmids used in this study. Figure S1. Growth curve of the wild-type ADP1 in mineral salts medium (MSM) supplemented with 10 g/L glucose and (A) furfural or (B) HMF at concentration of 0, 0.25, 0.5, 1, 1.5, and 2 g/L furfural or HMF. An overnight pre-culture was carried out in 5 ml MSM supplemented with 4 g/L glucose, 2 g/L casein amino acid, 0.1 g/L furfural, 0.1 g/L HMF, at 30 °C, 300 rpm. The main culture was carried out using 96-well plate, 200 µL cultivation, Spark multimode microplate reader (Tecan, Switzerland), at 30 °C. The experiment was repeated using independent biological triplicates, and the averages of the measurements, with error bars representing standard deviations are shown. Figure S2. Growth curve of wild-type ADP1 in SLH medium. An overnight pre-culture was carried out in 5 mL modified SLH medium, at 30 °C, 300 rpm. The main culture was carried out using 96-well plate, 200 µL cultivation, Spark multimode microplate reader (Tecan, Switzerland), at 30 °C. The experiment was repeated using independent biological triplicates, and the averages of the measurements, with error bars representing standard deviations are shown. Figure S3. Growth curve of wild-type ADP1 in MSM supplemented with 50 mM lactate as the sole carbon source. An overnight pre-culture was carried in 5 mL MSM supplemented with 10 g/L glucose, 2 g/L casein amino acid, at 30 °C, 300 rpm. The main culture was carried out using 96-well plate, 200 µL cultivation, Spark multimode microplate reader (Tecan, Switzerland), at 30 °C. The experiment was repeated using independent biological triplicates, and the averages of the measurements, with error bars representing standard deviations are shown. Figure S4. Visualization of WE accumulation of strains ADP1-ISx and ASA714 using thin layer chromatography (TLC). The cultivations were carried out at 30 °C, 300 rpm, using 100 mL Erlenmeyer flask with 10 mL MSM supplemented with 200 mM glucose, 300 rpm. The lipid extraction and the semi-quantitative analysis of WEs by TLC were done as described by Santala et al. Biomass samples were normalized to an OD600 of 20 of which 30 µL samples were applied on the TLC plate. TLC was carried out using Glass HPTLC Silica Gel 60F254 plates (Merck, USA) with a mobile phase of hexane: diethyl ether: acetic acid 90: 15: 1 (v/v/v). Wax esters on the TLC plates were visualized using iodine staining and using jojoba oil as the standard. Figure S5. Growth of ASA714 in SLH medium at varying lactate concentrations. The experiment was repeated using independent biological triplicates. An overnight pre-culture was carried in 5 mL SLH medium at 30 °C, 300 rpm. The main culture was carried out using 96-well plate, 200 µL cultures with SLH medium supplemented with 0–18 g/L lactate, Spark multimode microplate reader (Tecan, Switzerland), at 30 °C. The averages of the measurements, with error bars representing standard deviations are shown. Error bars are present but may be obscured by data symbols. Figure S6. (A) Growth profiles of the XXX strain in YP media containing 20 g/L glucose, 10 g/L glucose and 10 g/L xylose or 20 g/L xylose for 24 h. (B) Maximum ethanol concentration in each media after 9 h fermentation. The experiment was repeated using independent biological triplicates, and the averages of the measurements, with error bars representing standard deviations are shown. Figure S7. Growth profiles of the XXX strain in YPD medium supplemented with lactic acid at concentrations of 0, 5, 10, 30, 50, 100, 250, 500, 750, and 1000 mM lactic acid for 72 h. The experiment was repeated using independent biological triplicates, and the averages of the measurements, with error bars representing standard deviations are shown. Figure S8. Fermentation kinetics of LX3 in (A) YPD or (B) YPX media for 32 h. The experiment was repeated using independent biological triplicates. The averages of the measurements, with error bars representing standard deviations are shown. Figure S9. Fermentation kinetics including profiles of (A) cell growth, (B) sugar consumption, (C) lactate, (D) acetate, and (E) ethanol production of LX3 monoculture and coculture of LX3 and ADP1 at inoculation ratio of 1:2, 1:1, and 2:1. Fermentations were carried out in a 3.5 L bioreactor with 1 L SLH medium in 30 °C and pH was maintained at 6.0 by 2 M HCl and 8% (v/v) NH4OH solutions. Figure S10. Fermentation kinetics of (A) 1st, (B) 2nd, (C) 3rd, and (D) 4th LX3 monoculture in a bioreactor. Fermentations were carried out in a 3.5 L bioreactor with 1 L SLH medium at 30 °C and pH was maintained at 7.0 by 2 M HCl and 8% (v/v) NH4OH solutions. Figure S11. Fermentation kinetics of (A) 1st, (B) 2nd, (C) 3rd, and (D) 4th LX3 and ASA714 coculture in a bioreactor. Fermentations were carried out in a 3.5 L bioreactor with 1 L SLH medium at 30 °C and pH was maintained at 7.0 by 2 M HCl and 8% (v/v) NH4OH solutions. Figure S12. Growth and consumption of carbon sources by ASA714 in MSM supplemented with 8 g/L ethanol, 2.6 g/L acetate, and 2 g/L glucose. The cultivations were carried out using 100 mL Erlenmeyer flask with 10 mL cultivations, at 30 °C and 200 rpm. The experiment was repeated using independent biological replicates, and the averages of the measurements, with error bars representing standard deviations are shown.