title: Illumina HiSeq 4000 paired end sequencing; RNA-seq of wild-type Salmonella Typhimurium SL1344 and its AcrB D408A mutant
design: RNA-seq of wild-type Salmonella Typhimurium SL1344 and its AcrB D408A mutant
library_construction_protocol: Strains were grown in MOPS minimal medium, supplemented with histidine, to an OD600 of 0.6. All strains were incubated at 37°C, shaking at 180 rpm. Frozen bacteria pellets obtained from 3 mL of culture were used for extraction of total RNA. The Promega SV Total RNA Isolation kit was used according to manufacturer's instructions. Samples were heated at 70°C for 3 minutes and centrifuged for 10 minutes at full speed after addition of the RNA dilution buffer. RNA was eluted in water and 1 uL of an RNAse inhibitor was added to the extracted RNA. The library construction was done by BGI, Hong Kong. rRNA was depleted with Ribo-Zero Magnetic kit. cDNA was generated with First Strand Master Mix and Superscript II. Second Strand Master Mix was used consecutively. cDNA was combined with End Repair Mix and purified with Ampure XP Beads. Poly-A tails were added with A-Tailing Mix.