Accel ngs methyl seq dna library prep kit

WGBS libraries were prepared from 500 ng of genomic DNA. Isolated DNA (500 ng) from E8.5 mouse embryos (QIAamp DNA mini kit, 51304 or AllPrep DNA/RNA Micro Kit, Qiagen, 82084) and spiked-in lambda DNA (1.25 ng) (New England Biolabs) was sheared using the M220 ultrasound sonicator (Covaris) to an average genomic size of 350 bp. 200 ng of sheared DNA was bisulfite-converted using the EZ DNA Methylation-Gold Kit (Zymo Research, D5005) using manufacturer's instructions. Up to 100 ng of genomic DNA was then used to generate WGBS libraries using the Accel-NGS Methyl-Seq DNA Library prep kit (Swift Biosciences, 30024) according to manufacturer's instructions and 13 cycles of PCR for final library amplification. Final concentration of libraries were determined using the Qubit 1X dsDNA High Sensitivity Assay Kit (Invitrogen, Q33230) and final library sizes were assessed by agarose gel electrophoresis. Libraries were then sequenced in paired-end using the NovaSeq 6000 at the Center for Applied Genomics operated by the SickKids Research Institute.

Genomic DNA (500 ng) with spiked-in lambda DNA (1.25 ng) (NEB) was sheared using an M220 ultrasound sonicator (Covaris) to an average size of 350 bp. The size of the DNA fragments was confirmed by electrophoresis on a 1.5% agarose gel. 200 ng of sheared DNA was subjected to bisulfite conversion using EZ DNA Methylation-Gold Kit (Zymo Research) as per manufacturer’s instructions. Libraries were prepared using 50–100 ng of bisulfite-converted DNA and the Accel-NGS Methyl-Seq DNA Library prep kit (Swift Biosciences) according to manufacturer's instructions, with 8–11 cycles of PCR-amplification. The integrity of the libraries was assessed using agarose gel electrophoresis.
Libraries were sequenced on an Illumina HiSeq and NovaSeq 6000 instruments at the Michael Smith Genome Sciences Centre at BC Cancer and the Centre for Applied Genomics (SickKids), respectively.