Aqueous cell wall samples (200 µL) were applied over potassium bromide disks and dried at 50 ºC. Their infrared spectra were recorded by a Perkin-Elmer instrument with scanning range of 700–4200 cm−1 and resolution of 1 cm−1. RNA isolation and microarray hybridization Transcriptomic profiles of both M145 and INB201 strains were S. coelicolor determined using the data from the temporal series TS1 and TS3 of the ERA-NET SysMO project STREAM (GEO accessions GSE18489 and GSE31068). Experiments were performed in 3 liter fermentors as described by Nielset et al. (2010) and Thomas et al. (2012). For the transcriptomic analyses performed in this study, RNA samples from the liquid cultures were isolated after stabilization of 1 volume of culture with 2 volumes of RNA (Qiagen). The samples were treated with lysozyme (15 Protect Bacteria Reagent mg/mL), buffer RLT (Qiagen) and transferred to tubes (MP Lysing Matrix B Biochemicals) for mechanical disruption at homogenizer (speed 6.5, 30 sec, Fast prep Thermo Savant). The lysates were extracted with phenol-CIA and RNA was isolated from each sample using the (Qiagen), including the recommended RNeasy Mini Kit column DNase digestion to eliminate chromosomal DNA contamination. The final elution step was carried out with 50 µL of nuclease-free water. RNA samples from the TSA cultures were purified as follows: the cells were carefully taken from the plates with a small scraper (previously disinfected with a NaOH-SDS solution and commercial purchased from Ambion) and immediately put in a FalcomTM tube RNaseZap containing glass balls and 5 mL of Vigorous vortexing RNA Protect Bacteria reagent. was carried out to provide disaggregation of the cells from the agar remnants and
Aqueous cell wall samples (200 µL) were applied over potassium bromide disks and dried at 50 ºC. Their infrared spectra were recorded by a Perkin-Elmer instrument with scanning range of 700–4200 cm−1 and resolution of 1 cm−1. RNA isolation and microarray hybridization Transcriptomic profiles of both S. coelicolor M145 and INB201 strains were determined using the data from the temporal series TS1 and TS3 of the ERA-NET SysMO project STREAM (GEO accessions GSE18489 and GSE31068). Experiments were performed in 3 liter fermentors as described by Nielset et al. (2010) and Thomas et al. (2012). For the transcriptomic analyses performed in this study, RNA samples from the liquid cultures were isolated after stabilization of 1 volume of culture with 2 volumes of RNA Protect Bacteria Reagent (Qiagen). The samples were treated with lysozyme (15 mg/mL), buffer RLT (Qiagen) and transferred to Lysing Matrix B tubes (MP Biochemicals) for mechanical disruption at Fast prep homogenizer (speed 6.5, 30 sec, Thermo Savant). The lysates were extracted with phenol-CIA and RNA was isolated from each sample using the RNeasy Mini Kit (Qiagen), including the recommended DNase digestion to eliminate chromosomal DNA contamination. The final elution step was carried out with 50 µL of nuclease-free water. RNA samples from the TSA cultures were purified as follows: the cells were carefully taken from the plates with a small scraper (previously disinfected with a NaOH-SDS solution and commercial RNaseZap purchased from Ambion) and immediately put in a FalcomTM tube containing glass balls and 5 mL of RNA Protect Bacteria reagent. Vigorous vortexing was carried out to provide disaggregation of the cells from the agar remnants and column
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