GSM8230855: dTAG-SPT5-HCT116, muR1_3, Rep2; Homo sapiens; OTHER
About 2 million freshly prepared cell nuclei were used for one CUT&RUN experiment. The procedures were described in the manual of CUT&RUN kit (Epicypher). For DRB RNAPII CUT&RUN, dTAG-SPT5 cells grown on 100 mm dish were first treated with 125 nM dTAGv-1 for 4 hours, then followed by adding 100 μM DRB (5,6-dichloro-1-β-D-ribofuranosyl-1H-benzimidazole) for another 3 hours. DRB were washed out and cells were incubated in the fresh medium with 125 nM dTAGv-1 at 37 °C for the indicated time course. Cells were collected in the medium containing 1 µg/ml of actinomycin D on ice. Cell nuclei were extracted and proceeded with procedures above. Spike-In DNA (E.coli) (Epicypher) were added to each sample and DNA libraries were prepared with NEBnext Ultra II DNA library kit (New England Biolabs).