RNA-seq of coding RNA of Staphylococcus aureus COL wild type strain before and after 30 min of exposure to 176?µM neutrophil-derived oxidant hypothiocyanous acid (HOSCN) during exponential phase of an OD540 of 0.5 in LB medium
Staphylococcus aureus COL wild type strain was cultivated in LB medium in 3 biological replicates and harvested at an OD500 of 0.5 before (as control) and at 30 min after exposure to 1176?µM neutrophil-derived oxidant hypothiocyanous acid (HOSCN) stress. Cells were disrupted in 3 mM EDTA/ 200 mM NaCl lysis buffer with a Precellys24 ribolyzer followed by RNA isolation using the acid phenol extraction protocol as described. The RNA quality was approved by Trinean Xpose (Gentbrugge, Belgium) and the Agilent RNA Nano 6000 kit using an Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina (San Diego, CA, USA) was used to remove the rRNA. TruSeq Stranded mRNA Library Prep Kit from Illumina (San Diego, CA, USA) was applied to prepare the cDNA libraries. The cDNAs were sequenced paired end on an NextSeq 500  (San Diego, CA, USA) using 75 bp read length and a minimum sequencing depth of 8 million reads per library. The paired end cDNA reads were mapped to the Staphylococcus aureus COL genome sequence (accession number CP000046) using bowtie2 v2.2.7 (Langmead and Salzberg, 2012) with default settings for paired-end read mapping. All mapped sequence data were converted from SAM to BAM format with SAMtools v1.3 (Li et al., 2009) and imported to the software ReadXplorer v.2.2 (Hilker et al., 2016).