SRA

Francisella tularensis, the etiological agnet of Tularemia is a Category A select agent. Preparedness means for prompt proper antibiotic treatment are of need to prevent morbidity and mortality. While standard antimicrobial susceptibility tests are time consuming, the quantification of the changes in the expression levlels of specific mRNA markers folowing antibiotic exposure may enable a rapid determination of antimicrobial susceptibility. We submit transcriptomic data sets detailing global gene expression of F. tularensis following exposure to Doxycycline or Ciprofloxacin antibiotics. Overall design: F. tularensis SchuS4 colonies isolated on Cyctine Heart Agar (CHA) plates at 37oC were suspended in cation- adjusted Muller-Hinton broth, supplemented with 2% IsoVitaleX and 3 µM hematin (HLMHI) growth medium, to 5x105 CFU/ml. Suspended bacteria (200 ml) were grown for 2h (37oC, 200 rpm) to adjust the culture to the growth media. 50 ml of the adjusted bacterial culture were transferred into each of three 250 ml Erlenmeyer flasks and incubated in the presence of 0.5/1 x Minimal Inhibitory Concentration (MIC) of Ciprofloxacin/Doxycycline for 2 hours, followed by centrifugation (20 min., 4oC, 4,000xg). Bacterial pellets were resuspended in 1 ml cold PBS, transferred into Eppendorf tubes, centrifuged (5 min., 10K rpm at 400C), and pellets were frozen by liquid nitrogen and kept at -700C until RNA extraction. Growth control samples were grown under same conditions w/o antibiotic. The Ciprofloxacin / Doxycycline exposure experiments were done each by independent duplicated biological experiment. Total RNA purification and residual DNA elimination were done by the Rneasy Mini kit and Rnase-free Dnase kit (QUIGEN) according to the manufacturer's instructions. Experiments were performed using biosafety level 3 (BSL-3) containment and procedures. Sterile RNA samples were transferred to BSL-2 lab for following procedures. RNA concentration was determined by the NanoDrop ND-1000 spectrophotometer and RNA quality was determined by using the Agilent 2100 Bioanalyzer with the "Prokaryote Total RNA Pico" chip. All RNA samples showed RNA Integrity Number (RIN value) of >9. RNA samples were kept at -700C until used. Transcriptomic analysis was carried out by the RNA-seq method, at the Genome center, Columbia University.