GSM8335811: BT549_H3K27me3; Homo sapiens; OTHER
CUT&RUN: T47D and BT-549 cell lines were fixed in 0.1% formaldehyde for 1 minute. 5e5 cells were then washed and bound to activated ConA beads. The cell:bead complexes were then incubated with EZH2, H3K27me3 or H3K27ac antibodies overnight. Samples were then incubated with activated pAG-MNase and chromatin was subsequently released. 5 ng of purified DNA was used for multiplex library construction using the NEBNext Ultra II DNA Library Prep Kit. CUT&Tag: Nuclei were extracted from T47D cell line and were bound to activated ConA beads. The nuclei:bead complexes were then incubated with H3K27ac antibody overnight. Sample was then incubated with secondary antibody followed by activated pAG-Tn5. Tagmented chromatin was released and PCR amplified using barcoded Illumina primers for multiplex library creation. DNA purification was carried out with the PCR cleanup kit (NEB, Monarch PCR & DNA Cleanup Kit, #T1030). Libraries were prepared by NEBNext Ultra II kit (NEB, #E7645L) with 5ng CUT&RUN DNA as input.