GSM9461210: WT_10uM PQ_Rep1; Escherichia coli str. K-12 substr. MG1655; RNA-Seq
Bacteria were grown in M9 minimal medium supplemented with 4 g/L glucose, with respective paraquat concentration, till their mid-exponential phase. The culture was harvested and immediately processed for RNA isolation. The cells were dissolved in TRIzol and incubated at 65oC for 15 min. RNA was obtained in an aqueous layer upon the addition of chloroform, which was then precipitated using chilled isopropanol. This RNA pellet was washed with alcohol and finally resuspended in RNase-free water. RNA was quantified using a nano spectrophotometer before being frozen at -80°C for later use. RNA sequencing libraries were prepared using the QIAseq FastSelect rRNA depletion system (5S/16S/23S) in combination with the QIAseq RNA library preparation enzymes and buffers, following the manufacturer's protocols. After library construction, final library concentrations were quantified using the Qubit dsDNA High Sensitivity assay, and library size distribution and quality were assessed with the Agilent high-sensitivity NGS fragment analysis system. Libraries were normalized and pooled at equimolar concentrations prior to sequencing, and the pooled libraries were sequenced on an Illumina NextSeq 2000 platform.