GSM8442152: HUDEP-2, Bcl11a enhancer KO, diff, clone 2; Homo sapiens; OTHER
CUT&RUN was performed using the CUT&RUN Kit from Cell Signaling (catalog no. 86652S) according to the manufacturer's instructions. Briefly, 0.5 million cells were harvested and washed twice in 1× wash buffer at room temperature and bound to 15 μl of Concanavalin A beads. Primary antibody incubation was performed in PCR tubes for 2 h at 4 °C with rotation. H3K4me3 antibody supplied by the kit was used (1:xyz). After two washes, pAG-MNase was added and incubated for 1 h at 4 °C with rotation. After three washes, cells were resuspended in 1× wash buffer and pAG-MNase digestion was activated by adding 2 mM CaCl2. Samples were incubated on ice for 45 min, then 2× stop buffer was added to end the reaction. The samples were incubated at 37 °C for 10 min to release the captured chromatin fragments The sequencing libraries were prepared using the NEBNext Ultra II DNA Library (New England Biolabs, catalog no. E7645) with 15–30 ng of input DNA.