GSM8722934: R70H mNPC, Histone H2B ubiquitination, MA, Replicate 2; Mus musculus; OTHER
CUT&RUN assays were performed using the CUTANA kit following the manufacturer's protocol (EpiCypher, 14-1048). Briefly, mESCs Ring1B WT and R70H mutants were trypsinized (0.05%) to obtain a single-cell suspension followed by mild crosslinking with 0.1% formaldehyde for 1 minute. A total of 2 × 10⁵ cells were incubated with ConA beads, followed by overnight incubation with antibodies against HA (EpiCypher, 13-2010), Flag (EpiCypher, 13-2031), H3K27me3 (Active Motif, 39155), H2AK119ub (Cell Signaling D27C4), and IgG (EpiCypher, 13-0042). E. coli was used as a spike-in control for all samples. Additionally, the SNAP-CUTANA™ K-MetStat Panel (EpiCypher, 19-1002) was included as an additional spike-in control for IgG and H3K27me3. Two biological replicates were used per condition.