PacBio RSII
Long insert libraries were prepared as recommended by Pacific Biosciences guidelines for preparing size-selected 20Kb SMRTbellTM templates. Covaris g-TubesTM were used for shearing 10ug genomic DNA to fragments sizes of 10-25Kb. The PacBio SMRTbellTM library was size selected for fragments larger than 9Kb with the SAGE BluePippinTM devise. A second large insert library was prepared as described, but shearing of genomic DNA to 40Kb fragments was done with the MegaRuptorTM device (Diagenode) and this PacBio SMRTbellTM library was size selected for fragments larger than 10Kb. A total of 205 SMRT cells were sequenced on the PacBio RSII instrument making use of P4 polymerase and C2 sequencing chemistry. Movie length was 3 hours for all SMRT cells.