GSM2545744: Wild-type in vivo, replicate 1; Clostridium perfringens; RNA-Seq
Total RNA was then isolated using TRIzol reagent. The RNA aliquot was treated with Turbo DNase (Ambion) at 37°C for 30-60 min to eliminate DNA contamination. Contaminating host RNA was depleted from the sample using a MICROBEnrichTM kit (Ambion) in accordance with the manufacturer’s instructions.  A total of 5 μg of pooled RNA was subjected to repeated cycles of bacterial RNA enrichment to ensure the almost complete depletion of the host RNA. Next, bacterial ribosomal RNA (rRNA) was depleted from the preparation using a Ribo-Zero rRNA removal kit (Epicentre) as per the manufacturer’s instructions.  Libraries for bacterial RNA-seq were then prepared using an Illumina TruSeq RNA sample preparation kit v2.  RNA-seq analysis of the enriched RNA samples from the in vivo- and in vitro-derived cells was carried out on Illumina Hi-seq and Mi-seq instruments, respectively.