RNA-Seq of Chromobacterium violaceum: fur mutant; Biological replicate 3
The fur mutant was grown in LB medium (15 ml) at 37C with shaking until exponential phase (OD600nm 1). After centrifugation at 20 000 x g for 1 minute of 3 ml culture, total RNA from bacteria was extracted with 1 ml TRIzol (Ambion), followed by incubation for 10 minutes at 65C with 0.2 volumes of chloroform. RNA was purified with the Direct-zol RNA Miniprep Plus kit (Zymo Research). The analysis of RNA integrity was performed by running the samples on a denaturing agarose gel and in an Agilent 2100 Bioanalyzer (Agilent) with a Prokaryote Total RNA Nano kit. Depletion of ribosomal RNA (rRNA) and preparation of stranded-specific libraries were performed with the Illumina Stranded Total RNA, with Ribo-Zero Plus kit. The libraries were sequenced using the NextSeq 500/550 Mid Output v2 kit (150 cycles, 2x100) in an Illumina NextSeq2000 equipment, at NGS Solucoes Genomicas facility, Piracicaba, Brazil. The following sequences may be used to trim adapters. Read 1: AGATCGGAAGAGCACACGTCTGAACTCCAGTCA; Read 2: AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT.