After incubation, 0.5 mL of liquid media were taken from each culture medium and oxalate concentration in the samples was analyzed with a spectrophotometer (
Genesys 10S UV-vis, Thermo Scientific Inc., Waltham, MA, USA) at 590 nm as specified by the instruction manual (
Enzytec Oxalic acid, R-Biopharm AG). Samples were diluted 100-fold with distilled water before being used in the assay as they showed very high activities.
To determine any further changes in the protein secretions of
R.
placenta 45 exposed to the different amended media after incubation, 0.2 mL of liquid media was taken from each culture medium. Protein extracts from the control, CuCO
3·Cu(OH)
2, CuSO
4, MCA liquid media -with and without TTM- plus markers (BIO-RAD) were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using 10% gels loaded with 5 μL of each liquid culture sample and the marker. Gels were silver stained with the Dodeca
Silver Stain kit according to the instruction manual (BIO-RAD).
Civardi C., Schubert M., Fey A., Wick P, & Schwarze F.W. (2015). Micronized Copper Wood Preservatives: Efficacy of Ion, Nano, and Bulk Copper against the Brown Rot Fungus Rhodonia placenta. PLoS ONE, 10(11), e0142578.