Ion Xpress Fragment Library Prep

Library preparation from ChIP-isolated DNA was performed using the Ion Xpress™ Plus Fragment Kit and the Ion Xpress™ Barcode Adapters 1–16 Kit (both from ThermoFischer Scientific Inc., Grand Island, NE, USA). The protocol for 200 bp read-length libraries and for 50–100 ng input DNA was employed with some modifications. Due to the small amount of input DNA (10 ng), and to avoid adapter concatamerization, 1 μL of P1 adapter and IonXpress™ Barcodes was used instead of 2 μL. Adapter ligation and dual bead size selection for 200 bp read length fragments was performed using AMPure XP beads, followed by separation on magnetic rack, two ethanol washes and a final elution of DNA fragments in 25 μL Low TE. The size selected libraries were then amplified for 16 cycles. The quality was assessed on the Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA) using a High Sensitivity DNA ChIP (Agilent Technologies, Santa Clara, CA, USA), and quantified via qPCR using the KAPA Library Quantification Kit for Ion Torrent™ [54 (link)]. Template preparation and chip loading prior to sequencing was performed on the Ion Chef system using the Ion 540™ Chef kit and an Ion 540™ chip. The loaded chip was then sequenced on an Ion GeneStudio S5 sequencer.