title: GSM6283072: WGBSseq_hek293_qko_mbd; Homo sapiens; Bisulfite-Seq
library_construction_protocol: Nuclei of neurons (day 8) were isolated as in Grand et al.(Grand et al. 2021) and sorted by flow cytometry (BD FACS Aria III). Genomic DNA was isolated (QIAamp DNA Micro Kit, catalog number 56304) from  mouse ES cells or sorted nuclei of neurons and 1 ug was fragmented (Covaris S220) to an average size of ~ 300 bp. HEK293 genomic DNA was isolated with QIAamp DNA mini kit (Qiagen catalog number 51306) and fragmented with Covaris S220. Libraries of mouse DNA were prepared according to manufacturer's instructions: Adapter ligation was performed using NEBNext Ultra II DNA Library Prep Kit (catalog number E7645L) with methylated adapters (NEBNext, catalog number E7535S), bisulfite treated (EZ DNA Methylation-Gold™ Kit, Zymo catalog number D5006) and indexed (NEBNext® Multiplex Oligos for Illumina) using 11 cycles in the PCR reaction (KAPA HiFi HotStart Uracil+ ReadyMix, Roche catalog number 07959052001). For HEK293 DNA, 500 ng of fragmented DNA were used for library preparation (NEBNext Ultra DNA Library Prep Kit, NEB catalog number E7370) with methylated adapters (NEBNext, NEB catalog number E7535S) and bisulfite treated with EZ DNA methylation-lightning Kit (Zymo Research catalog number D5046). Final PCR amplification was performed using KAPA HiFi HotStart Uracil+ ReadyMix PCR Kit (Roche catalog number 07959052001) with 12 cycles of amplification.