RNA-Seq Analysis of E. coli Transcriptome

RNA sequencing (RNA-Seq) was performed and analyzed as recently described (18 (link)). Briefly, overnight cultures were diluted to OD₆₀₀ = 0.1, grown for 1 h, and treated or not with 10 μM TAT-RasGAP_317-326 for 1 h. RNA was extracted with RNeasy Plus minikit (Qiagen), and any remaining DNA was removed using DNA-free DNA Removal Kit (Invitrogen, Carlsbad, CA). Integrity of the resulting RNA samples was verified by Standard Sensitivity RNA Analysis kit (Advanced Analytical, Ankeny, IA) with the Fragment Analyser Automated CE System (Labgene Scientific, Châtel-Saint-Denis, Switzerland). Preparation of the libraries and Illumina HiSeq platform (1 × 50 bp) sequencing were performed by Fasteris (Plan-les-Ouates, Switzerland). Reads were then trimmed and mapped to genome of E. coli K-12 MG1655. Normalized expression values were calculated as reads per kilobase of transcript per million mapped reads (RPKM) with edgeR (50 (link)).