GSM7956977: guidedNOMe-seq in Adnp-FKBP12(F36V) mES cells after 4 hours dTAG13 treatment, i.e. ADNP depletion, rep2; Mus musculus; OTHER
0.8*10e6 cells were harvested at the indicated time-points, washed and nuclei were treated with M.CviPI for methylation foot-printing. Then cells were lysed and DNA was isolated. 120ng DNA was dephosphorylated with rSAP and digested by Cas9 loaded with an in vitro transcribed guideRNA pool for the regions of interest.The digested DNA was used togenerate an NGS library. For a detailed protocol, please see Schwaiger et al, 2024. Libraries were generated using the NEBNext Ultra II DNA Library Prep Kit and xGen Methyl UDI-UMI (IDT) premethylated adapters. The library was bisulfite treated with the EZ DNA Methylation-Direct Kit, amplified using the KAPA HiFi Uracil+ (Roche) Kit, size selected with Ampure beads (targeting 300-600bp)and sequenced on an Illumina MISeq instrument in 600bp paired-end mode. For a detailed protocol, please see Schwaiger et al, 2024.