title: GSM4223305: Salmonella Derby ER278::hilD_v - rep1; Salmonella enterica subsp. enterica serovar Derby; RNA-Seq
library_construction_protocol: 0.6 mL of each culture (about 1.2 x 109 CFU/mL) was centrifuged at 16,000g for 2 minutes and pellets were frozen in a dry ice ethanol bath. Cells, resuspended in Lysis Buffer, were processed in a FastPrep-24 Instrument for 90 seconds, at 6.0 m/s, for cell disruption. RNA extraction was performed by FastRNA SPIN Kit for Microbes (MP Biomedicals) according to the manufacturer instruction. Trace DNA was removed by DNase digestion with TURBO DNA-free kit (Invitrogen Ambion), using rigorous treatment. Complete loss of genomic DNA after DNase treatment was verified by PCR on RNA samples using rpoD_FW and rpoD_RV primers. RNA samples were then precipitated for purification. RNA quality was assessed using a 2100 Bioanalyzer (Agilent) according to Agilent RNA 6000 Nano Kit protocol. Each sample was then retro-transcribed by SuperScript VILO cDNA Synthesis Kit (Invitrogen). In order to obtain double-strand cDNA for libraries preparation, single-strand cDNA samples were purified using 1.8X Agencourt AMPure XP Beads (Beckman Coulter) and the second cDNA strand was synthesized using NEBNext Ultra II Non-Directional RNA Second Strand Synthesis. A second step of beads-based purification was performed before libraries preparation. Libraries were prepared using Nextera XT V2 kit according to the reference guide.
attributes.tag: GEO Accession
attributes.value: GSM4223305