GSM3142243: N2_60_1; Clostridioides difficile; RNA-Seq
Total bacterial RNA was isolated from bacterial cell pellets, from three independent experiments, using acid phenol and glass beads. DNA was digested using TURBO DNase (Ambion) and the DNA-free RNA was purified with phenol:chlo­roform:isoamylalcohol (25:24:1). Remaining traces of phenol were removed by washing the samples twice with chloroform:isoamylalcohol (24:1). RNA integrity was assessed using the Agilent RNA 6000 Nano Kit on the Agilent 2100 Bioanalyzer. To 2 µg of total RNA 1 µL of 1:10 diluted ERCC ExFold RNA Spike-In Mix 1 or 2 (Ambion) was added. rRNA was depleted using the Ribo-Zero rRNA Removal Kit (Gram-Positive Bacteria) as recommended by the manufacturer (Epicentre). Strand-specific barcoded cDNA libraries were prepared using the ScriptSeq v2 RNA-Seq Library Preparation Kit as recommended by the manufacturer (Epicentre). Library quality was validated using Agilent 2100 Bioanalyzer (Agilent Technologies) following the manufacturer's instruction.
GEO Accession
GSM3142243