Genome sequence of Pseudomonas aeruginosa ST664 clinical isolate
The quality of the genomic DNA samples was assessed by running 150ng of DNA on a 0.6% pulsed-field agarose gel, stained with SYBR Safe (Invitrogen). Ten micrograms of DNA were then sheared to a size range of 10-40kb using a Covaris g-TUBE. The fragment distribution of the sheared samples was again validated by pulsed-field gel electrophoresis. The sheared DNA was then purified with 0.45X AMPure PB beads (Pacific Biosciences) according to the manufacturers recommendation. Library preparation was performed by following Pacific Biosciences 20kb SMRTbell Template Preparation Protocol, using 5ug of the sheared DNA as input. After library preparation, libraries were assessed on an Agilent DNA 12000 bioanalyzer chip to determine the optimal cut-off for size selection. The libraries were then size-selected on a Sage Science Blue Pippin instrument, using a dye-free 0.75% agarose cassette and 15kb as the cut-off. Each sample was then sequenced in 1 SMRTcell on a Pacific Biosciences RSII single-molecule sequencing platform at a loading concentrations of 0.25nM.