The Ion Torrent™ Personal Genome Machine™ (PGM™) is revolutionizing the sequencing world with its “Ion Semiconductor Sequencing”. However, its manual library preparation workflow presents a bottleneck. The Apollo 324™ is a benchtop instrument that fully automates the key steps of the Ion Fragment Library Preparation processing using the PrepXTM PGM Library preparation kit. After a 15-minute setup of reagents on the instrument, the end repair, adapter ligation and size selection are all performed with walk-away operation in just under 3.5 hours. The basic workflow is as follows:
Sample Preparation: Raw DNA samples are sheared using the recommended Covaris® settings to provide a 1 μg (15 mL) sheared DNA sample with a broad size distribution centered at around 200 bp.
Library Preparation: The sheared DNA, reagents, disposables, and AMPure® XP beads are loaded onto the Apollo 324 instrument work surface. The end repair, adapter ligation and size selection are all performed in less than 3.5 hours using AMPure XP beads for both clean-up and size selection. As many as eight library samples can be prepared per run, and two runs can be performed per day.
Library Fragment Quality Assessment: Proper size selection is confirmed by running an Agilent® High Sensitivity Chip. The initial version of the protocol prepares a library fragment with 80% of the total sample between 150-250 bp and an average size of 195 bp, eliminating the need for time-consuming gel size selection. Nick translation and enrichment PCR are typically performed. The resulting DNA library has comparable sequencing results to libraries prepared using the Caliper LabChip® XT system.