GSM8140744: RHA1 on 30 mM glycolate, mid-log phase, rep3; Rhodococcus jostii; RNA-Seq
The cell pellets were suspended in 2 ml TRIzol™ (Thermo Fisher Scientific) and stored at -80 °C. One ml of cells in TRIzol™ were transferred to 2 ml screw cap tubes with glass-beads and lysed by using a MP Biochemical bead-beater (three rounds for 45 sec at power 6.5) with a 10 min incubation step at 60 °C in between. RNA extraction was then proceeded according to the manufacturer's instructions and treated with TURBO™ DNase (Thermo Fisher Scientific). Quality and quantity of nucleic acids were assessed using 1% (w/v) agarose gel electrophoresis and Qubit fluorometric quantitation (Thermo Fisher Scientific), prior to storage at −80°C. Genomic DNA contamination was assessed by using RT-qPCR and probes for the sigA marker gene. One μg of rRNA was depleted using the NEBNext® rRNA Depletion Kit (Bacterial) and the libraries were prepped with NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina®. The quality of the rRNA-depleted samples and the libraries were checked by using a Bioanalyzer 2100 and the RNA 6000 Nano kit (Agilent).