Table 3.
Comparison of C. glutamicum ATCC 13032 and MB001a
| Type of expt | Conditions | Resultc |
|---|---|---|
| Growth | ||
| Different C sourcesb; see Fig. S1 in the supplemental material | CGXII minimal medium with glucose, gluconate, acetate, or lactate as carbon source | x |
| Bioreactor scale | CGXII minimal medium with 56 mM glucose, 1-liter bioreactor, 200–1,200 rpm; DO, 30%; pH 7.0 | x |
| Microfluidic scale | CGXII minimal medium with 222 mM glucose, array of picoliter bioreactors, continuous medium supply, 100-nl/min flow rate | x |
| Stress resistance | ||
| Iron excessb | CGXII minimal medium with 56 mM glucose, 25–3,125 μM FeSO4 | x |
| Osmotic stressb | CGXII minimal medium with 56 mM glucose, 1–2,000 mM NaCl | x |
| Phosphate starvationb | CGXII minimal medium with 56 mM glucose, 13–0.65 mM phosphate | x |
| Mitomycin C treatment; see Fig. 2 | CGXII minimal medium with 111 mM glucose and 1 μM mitomycin C | Growth rates, ATCC 13032 vs MB001: 7–12 h, 0.15 ± 0.01 vs 0.17 ± 0.00; 15–17 h, 0.18 ± 0.02 vs 0.27 ± 0.01 |
| Agar diffusion assay; see Fig. S2 | 6 M NaOH, 6 M HCl, 30% H2O2, ethambutol, penicillin, vancomycin, Tween 40 | x |
| Plate assays; see Fig. S3 | UV radiation (0.5–5 min), mitomycin C added (10–1,000 nM), iron starvation (0 μM), iron excess (100 μM), heat shock (42°C, 30–60 min), osmotic stress (1 M NaCl) | x |
| Heterologous protein production | ||
| Production of eYFPb (Western blot); see Fig. 4 | Plasmid pEKEx2-eyfp, CGXII minimal medium with 56 mM glucose and 1 mM IPTG | 40% increased eYFP signal and increased amount of eYFP protein in strain MB001 |
| Plasmid copy no.b (qPCR); see Fig. 4C | Plasmid pEKEx2-eyfp, CGXII minimal medium with 56 mM glucose and 1 mM IPTG | 2- to 3-fold-increased plasmid copy no. in MB001 |
| Other | ||
| Transcriptomics, ATCC 13032 ΔCGP3 vs ATCC 13032 (see Table S3) | CGXII minimal medium with 111 mM glucose, harvested in exponential growth phase at OD600 of 5 | x |
| Transformation efficiency; see Fig. 3 | Tested for integrative and replicative plasmids | 2- to 3-fold increase for integrative plasmid and 5- to 8-fold increase for replicative plasmid in strain MB001 |
a
Overview of all experiments for comparison of wild-type ATCC 13032 and the prophage-free strain MB001 performed in this study.
b
Cultivation in the BioLector system in 48-well Flowerplates (m2p-labs, Germany).
c
x, no significant difference was observed under the chosen conditions.