GSM9229786: NPM1c CUT&Tag, OCI-AML3 cells, siControl, Day3, rep1; Homo sapiens; OTHER
1 × 10^5 OCI-AML3 cells were collected and washed once with 500 μl wash buffer before they were bound to ConA beads for 10 min at 25 °C. Cells were incubated with 1 mg of respective targeting antibodies at 4 °C overnight. 0.5 mg secondary antibody was added and incubated for 1 h at 25 °C. Then cells were washed three times with DIG wash buffer and incubated with 0.04 μM pA/G–Tnp for 1 h at 25 °C. Similarly, cells were washed three times with DIG 300 buffer, resuspended in tagmentation buffer and incubated at 37 °C for 1 h. Tagmentation was stopped by adding proteinase K, buffer LB and DNA extract beads. After incubation at 55 °C for 10 min, cells were plated on a magnet and unbound liquid was removed. Beads were gently rinsed twice with 80% ethanol and DNA was eluted with double-distilled water. libraries were barcoded using TruePrep™ Index Kit V2 for lllumina (Vazyme TD202) and amplified for 14 cycles. Final product was cleaned by VAHTS DNA Clean Beads at a 2 × ratio. Libraries were sequenced on a Novaseq 6000 in a 150 bp/150 bp Paired end run. An average of 20 × 106 paired reads was generated per sample.