Illumina HiSeq 2500 paired end sequencing; RNA-seq of E. coli strain UTI89 with isogenic mutants lacking QseB and PmrA following activation with ferric iron
RNA-seq of E. coli strain UTI89 with isogenic mutants lacking QseB and PmrA following activation with ferric iron
Samples were collected from cultures allowed to reach the mid-logarithmic growth phase in N-minimal media. At this time 100 uM ferric iron was added to the medium.  Samples were collected prior to addition of ferric iron and at 15 and 60 minutes post addition. 4 mL of cells were collected and pelleted at these times, flash frozen, and stored at -80C until RNA extraction. Strains were grown at 37C overnight shaking in lysogeny broth. The next day, strains were subcultured in fresh N-minimal media and allowed to reach mid-logarithmic growth.   Cell pellets were thawed from-80C storage and total RNA was extracted according to Qiagen RNeasy kit instructions.  VANTAGE (Vanderbilt Technologies for Advanced Genomics) performed mRNA enrichment using the Illumina Ribo-Zero kit and prepared cDNA libraries with the Illumina Tru-seq stranded mRNA sample prep kit.