2.1 Media and culture conditions
Plasmids and strains used in this study can be found in Table S1 in the
supplementary information. For cloning purposes, Escherichia coliand P. taiwanensis cells were cultivated at 37 or 30 °C,
respectively, either in liquid LB medium containing 5 g
L−1 sodium chloride or on solid LB agar plates (with
1.5 % (w/v) agar). After conjugational mating procedures, Pseudomonads
were isolated on cetrimide agar (Sigma Aldrich) plates supplemented with
10 mL L−1 glycerol. Kanamycin (50 mg
L−1) or gentamicin (20 mg L−1) was
added to cultures or plates when necessary. Growth and production
experiments were performed in mineral salts medium (MSM) [9] with 20
mM glucose or 40 mM glycerol as sole carbon source.
In production experiments, liquid cultures of P. taiwanensis were
performed in MSM with 20 mM glucose or 40 mM glycerol without the
addition of antibiotics. Main cultures were inoculated at an
OD600 of ~0.2, from seed cultures grown
in MSM containing glucose. Batch production experiments were performed
in 500 mL Erlenmeyer flasks with a culture volume of 50 mL, cultivated
in a rotary shaker with a frequency of 200 rpm and a throw of 50 mm. Fed
batch fermentations were performed in DASbox® mini-bioreactors
(Eppendorf) according to the setup and procedure described in Otto et
al. [8].