Cultivation of Rhodobacter capsulatus Strains

Three Rhodobacter capsulatus strains were used in this study: rifampicin sensitive wild-type strain B10 (Wall et al., 1975 (link)), a rifampicin resistant derivative SB1003 (ATCC BAA-309) and an RcGTA overproducer strain DE442 (Ding et al., 2014 ; Fogg et al., 2012 (link)). All R. capsulatus cultures were grown at 30°C either aerated in the dark or in anoxic sealed tubes under constant illumination. Two growth media were used – YPS complex broth (0.3% w/v yeast extract, 0.3% w/v peptone, 2 mM MgCl₂, 2 mM CaCl₂) or RCV defined broth (10 mM potassium phosphate buffer, 0.4% w/v L-malic acid, 0.1% w/v (NH₄)₂SO₄, 0.020% w/v MgSO₄^.7H₂O, 0.0075% w/v CaCl₂^.2H₂O, 0.0012% w/v FeSO₄^.7H₂O, 0.0020% w/v Na₂EDTA, 0.0001% w/v thiamine hydrochloride. Plus 1 mL of trace element solution - 0.07% w/v H₃BO₃, 0.040% w/v MnSO₄^.H₂O, 0.019% w/v Na₂MoO₄^.2H₂O, 0.006% w/v ZnSO₄^.7H₂O, 0.001% w/v Cu(NO₃)^.3H₂O. The pH was adjusted to 6.8 with NaOH). For agar plates, 1.5% w/v agar was added to the above broth recipes. The E. coli S17-1 strain (DSM 9079), which contains chromosomally integrated tra genes, was used as a donor for all conjugations. NEB 10-beta Competent E. coli cells (New England Biolabs) were used for standard cloning and plasmid maintenance; T7 Express Competent E. coli cells (New England Biolabs) were used for overexpression of proteins for purification.