E. coli and B. subtilis growth and media conditions were similar to those described previously9 (link), 20 (link). Briefly, E. coli were grown on LB agar with an antibiotic as needed at 37°C overnight before being inoculated into LB or 2XYT liquid medium with an antibiotic as needed and grown at 37°C.
B. subtilis cultures were inoculated from single colonies and were grown in LB Lennox and S7 liquid media as indicated at 37°C. For the preparation of S7 growth media, we modified the Teknova MOPS Minimal Media kit (M2106) to the appropriate concentrations and supplemented with 0.1% w/v glutamic acid and 40 μg/mL L-tryptophan.
For FRET experiments, single colonies grown on LB Lennox agar overnight were inoculated into S7 media and grown in a roller drum to early log phase. For imaging purposes, the cultures were then concentrated via centrifugation (3000 rcf, 2 min) and resuspended in the original supernatant.
For antibiotic treatment experiments, 3ml cultures were grown to midlog (OD600~0.4) in S7 media before being split and treated with the antibiotic at the concentration indicated for 1.5 h. Antibiotic sensitivity experiments were performed using ETest® strips (Biomerieux) on S7 agar plates at 37°C overnight.
B. subtilis cultures were inoculated from single colonies and were grown in LB Lennox and S7 liquid media as indicated at 37°C. For the preparation of S7 growth media, we modified the Teknova MOPS Minimal Media kit (M2106) to the appropriate concentrations and supplemented with 0.1% w/v glutamic acid and 40 μg/mL L-tryptophan.
For FRET experiments, single colonies grown on LB Lennox agar overnight were inoculated into S7 media and grown in a roller drum to early log phase. For imaging purposes, the cultures were then concentrated via centrifugation (3000 rcf, 2 min) and resuspended in the original supernatant.
For antibiotic treatment experiments, 3ml cultures were grown to midlog (OD600~0.4) in S7 media before being split and treated with the antibiotic at the concentration indicated for 1.5 h. Antibiotic sensitivity experiments were performed using ETest® strips (Biomerieux) on S7 agar plates at 37°C overnight.
