GSM4145666: Control replicate 3; Staphylococcus aureus; RNA-Seq
S. aureus recovered from infected macrophages or from control samples were  Centrifuged at 5000g for 5 min and lysed using the following procedure: pellets were resuspended in  Tris-EDTA buffer containing freshly prepared 13 mg/mL lysozyme (Sigma) and 130 μg/mL lysostaphin (Sigma) for 30 min at room temperature. Resulting suspensions were processed for total RNA extraction with RNA extraction InviTrap Spin Universal RNA Mini Kit (Stratec) following the  manufacturer's instructions. Traces of contaminating genomic DNA were removed from samples by  treatment with TURBO DNase (Ambion) for 30 min at 37°C according to the manufacturer's  instructions. RNA purity was checked using a NanoDropM spectrophotometer (Thermo Fisher  Scientific). Total RNA from three independent replicates were checked on the Bioanalyser system (Agilent) for its quality and integrity. Ribosomal RNA depletion was performed using the Bacteria RiboZero kit (Illumina). From rRNA-depleted RNA, directional libraries were prepared using the TruSeq Stranded mRNA Sample preparation kit following the manufacturer's instructions (Illumina). Libraries were checked for quality on Bioanalyser DNA chips Bioanalyser (Agilent). Quantification was performed with the fluorescent-based quantitation Qubit dsDNA HS Assay Kit  (Thermo Fisher Scientific). Sequencing was performed as an SRM run (SR: Single Read, PE: Paired-end Reads, M: multiplexed samples) for 65 bp sequences on HiSeq 2500 Illumina sequencer (65 cycles). The multiplexing level was 6 samples per lane.
GEO Accession
GSM4145666