PacBio sequencing of E. coli o157:H7 FRIK804: Isolated from a calf in Wisconsin
gDNA from purified using the method Bacterial genomic DNA isolation using CTAB from the JGI protocol (version 3) by William S. Feil, Helene Feil, and A. Copeland. The gDNA sample was submitted to the University of Wisconsin-Milwaukee Great Lakes Genomic Center. A standard Pacific Biosciences large insert library was prepared by fragmenting DNA to approximately 20 kb using g-TUBEs (Covaris, Woburn, MA). Fragmented DNA was enzymatically repaired and ligated to a PacBio adapter to form the SMRTbell Template. Templates larger than 10kb were size selected using BluePippin (Sage Science, Beverly, MA). Templates were annealed to sequenced primer, bound to pilmerase (P6), and then bound to PacBio Mag-beads and SMRTcell sequenced using a RSII sequencer and C4 chemistry.