GSM9406918: U2OS_Control_IgG; Homo sapiens; OTHER
Sample preparation was done with CUTANA™ ChIC / CUT&RUN Kit (EpiCypher #14-1048). Cells (500,000) were fixed in culture medium supplemented with 0.1% formaldehyde for 1 min before quenching with 0.125 M glycine. Cells were then permeabilized, bound to concanavalin A magnetic beads and incubated overnight at 4 °C with primary antibodies using a 1:50 dilution. After overnight incubation and washing to remove excess antibodies, protein A/G-MNase was added to the antibody-bound beads and activated with calcium at 4 °C for 2 h to cleave target-bound genomic DNA. After quenching the cleavage reaction, the samples were reverse cross-linked under proteinase K treatment overnight at 55 °C. CUT&RUN DNA was isolated with column chromatography, and libraries were prepared using an NEBNext Ultra II DNA Library Prep kit for Illumina (New England BioLabs). Libraries were sequenced on an Illumina NovaSeq 6000 with paired-end 150-base pair reads.