Oxidative stress response in R. sphaeroides

R. sphaeroides strains (Table C in S1 File) were cultivated in malate minimal salt medium [14 (link)] at 32°C in the dark. Conditions of semi-aerobic (~25 μM dissolved oxygen) and aerobic growth (~180 μM dissolved oxygen) were accomplished as recently described [28 (link)]. Iron-limiting conditions were achieved as described in [15 (link)]. Stress experiments were conducted in exponential growth phase (OD₆₆₀ of 0.4). Singlet oxygen (¹O₂) was generated by the addition of 0.2 μM methylene blue (Sigma-Aldrich) in the presence of high light intensities (800 W m^-2 white light). Hydrogen peroxide (H₂O₂; Roth) and tert-butyl hydroperoxide (tBOOH; Sigma-Aldrich) were added in final concentrations of 1 mM and 300 μM, respectively. 250 μM of paraquat (PQ) were used for the generation of superoxide radicals (O₂^‒). Samples taken at the indicated time-points were rapidly cooled on ice and harvested by centrifugation at 10,000 x g in a chilled centrifuge.