title: Illumina MiSeq paired end sequencing; RNA-Seq of roseoflavin producer Streptomyces davaonensis reveals transcription of unusual high number of riboflavin biosynthetic genes (rib genes) in exponential and stationary growth phase
design: RNA-Seq of roseoflavin producer Streptomyces davaonensis reveals transcription of unusual high number of riboflavin biosynthetic genes (rib genes) in exponential and stationary growth phase
library_construction_protocol: S. davaonensis was grown in YS and cells were harvested in the exponential growth phase (after 18 h of growth) and from the stationary growth phase (after 48 h of growth). For each condition three independent cultures inoculated with the same amount of spores were analyzed.  S. davaonensis was grown in YS and cells were harvested in the exponential growth phase (after 18 h of growth) and from the stationary growth phase (after 48 h of growth). For each condition three independent cultures inoculated with the same amount of spores were analyzed.  Total RNA was isolated from three biological replicates using RNeasy Mini Kit along with a DNase Kit (both from Qiagen). Initially RNA quality was checked by Trinean Xpose (Gentbrugge,Belgium) and Agilent RNA Nano 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). Samples contaminated with DNA were treated with DNase (Qiagen), cleaned as described above and rechecked by Xpose and Agilent Bioanalyzer. Finally RNA was free of DNA with an RNA Integrity Number (RIN) > 9 and rRNA Ratio [23s / 16s] > 1.5. Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina (San Diego, CA, USA) was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was checked by Agilent RNA Pico 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). RNA was free of detectable rRNA. TruSeq Stranded mRNA Library Prep Kit from Illumina (San Diego, CA, USA) was used to prepare cDNA libraries.