GSM6742817: 1338over, DETA/NO, replicate 3; Mycobacterium tuberculosis; RNA-Seq
Bacterial cultures were rapidly cooled on ice, centrifuged, and total RNA was isolated by phenol-chloroform extraction after cell disruption with Bead Beater. After isolation, RNA was treated with Turbo DNase (Life Technologies, Carlsbad, CA, USA) to remove traces of genomic DNA, and purified with the RNeasy mini kit (Qiagen, Venlo, Netherlands). RNA samples were depleted of 16S and 23S rRNA using Ribonuclease H (NEB) and custom oligonucleotides according to the protocol published by Huang et al. [10.1093/nar/gkz1169]. Sequencing libraries were generated using the resulting ribosomal transcript-depleted RNA and NEBNext Ultra II Directional RNA Library Prep Kit (NEB) according to the manufacturers' protocol.