RNA-Seq of Enterococcus faecalis enriched colon content of gnotobiotic SIHUMI colonized mice (129Sv/Ev wild type)
E. faecalis cells in the colonic content of SIHUMI colonized mice (colonization period:16 weeks) were isolated by immunomagnetic separation. Total bacterial RNA was isolated and purified using the RNeasy MinEluteTM Cleanup kit. gDNA was eliminated using TURBO DNase. rRNA was depleted by using the RiboMinus Transcriptome Isolation Kit. RNA was fragmented using a Covaris E220 sonicator and dephosphorylated using RNA Antarctic phosphatase. The RNA fragments were recovered using the miRNeasy Mini kit followed by 5' phosphorylation using T4 polynucleotide kinase. Again, the miRNeasy Mini kit was used for recovery. The cDNA library was prepared as described in the TruSeq Small RNA Library Prep Reference Guide with the following exception. After adapter ligation and reverse transcription, fragments were size selected between 145bp and 300bp corresponding to an insert length between 18bp and 173bp.