RNAseq_Saureus_H2O2_vs_Control
Cultures of SA925 and SA1088 (5 mL) were grown overnight in TSB from single colonies. Samples were back diluted 1/1000 and grown with and without 1% H2O2 shaking at 37 C to an OD 600 of 1.0. RNA was extracted from each sample by resuspending each bacterial pellet in 1 mL TriZol and homogenizing in a BioSpec MiniBeadbeater 16 with 0.1 mm zirconium particles (BioSpec) followed by centrifuging lysate at 14,000 g for 3 minutes. Following centrifugation, supernatant was mixed with .5mL 95% ethanol and treated using the Direct-zol RNA Miniprep Plus Kit (Zymo Research) and treated with DNAse 1. RNA integrity was assessed with a 2100 Bioanalyzer and the Prokaryote Total RNA 6000 Pico Kit (Agilent). All samples had an RNA Integrity Number (RIN) >8. RNA concentration was quantified on Qubit using the RNA Broad Range assay (Invitrogen). Ribosomal RNA was depleted and libraries were constructed using the Illumina Stranded Total RNA Prep with Ribo-Zero Plus (Illumina) using 100 ng of RNA input.