95% pHW197-M : 5% pHW197-Mmut on Ion Torrent PGM
To construct the library, 100 ng was first sheared with an M220 focused-ultrasonicator (Covaris) set to obtain peak fragment lengths of 400-500 bp. After shearing, blunt ends were created using the end repair enzyme from the Ion Plus Fragment Library kit (Life Technologies). Next, the fragments were ligated to Ion Torrent PGM-compatible barcode adapters. Since the adaptors are not 5' phosphorylated, the nick repair polymerase in the kit repairs subsequently the nick on one strand at each ligation site, in order to minimize adaptor-dimer formation. We purified and simultaneously size-selected the adapter-ligated library using Agencourt AMPure XP bead sizing (Beckman Coulter).