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Agrobacterium containing the respective constructs were cultured in 5 mL LB medium containing respective antibiotics for 24 hr at 28°C.
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Twenty-four hours after transfection, the medium was replaced by medium containing a respective ligand and/or a testing compound.
The following day, the medium was replaced with medium containing the respective selection antibiotic (G418).
The medium containing the respective supplements was replaced every 2 days.
The experiments were repeated twice and medium was changed every second day to fresh medium containing the respective agents.
After reaching the stationary phase, cells were transferred into fresh medium containing the respective substrate to confirm growth.
Cells were also cultivated in the fermentor with acetate, octanal and octanoate under the same conditions but with a mineral salts medium containing the respective organic compounds.
After reaching the stationary phase, aliquots were transferred into fresh medium containing the respective substrate; if appropriate, repeated passages were performed to confirm growth.
For screening purposes and for maintenance of protein expression potent selection markers need to be used and producer cells have to be constantly cultivated in medium containing the respective selective agent.
Cells were seeded onto 6-well plates in their respective medium containing 3% (v/v) charcoal-stripped serum (CSS).
Cells were washed with cold PBS and resuspended with respective medium containing TMRE at a final concentration of 40 nM.
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