GSM6503406: PEDS0009, sgChr2, FLI1; Homo sapiens; OTHER
CUT & RUN (Cleavage Under Targets & Release Using Nuclease) was performed as previously described105-107 with slight modifications. Briefly, 500,000 cells per condition were bound to activated ConA Beads (EpiCypher 21-1401). Next, the ConA bead-cell mixture was resuspended in a cold antibody buffer and FLI-1-(ab133485; 1mcg per sample) antibody or 0.5 mcg H3K4me3 (EpiCyper 13-0041) as positive and 0.5 mcg  IgG (EpiCypher 13-0042) as negative control were  per sample added over night. Next, pAG-MNase (EpiCypher 15-1016) was added to each reaction to allow binding to the antibody-labeled chromatin. E.coli Spike-in DNA (EpiCypher 18-1401) was added following MNase activation. Subsequently, targeted chromatin was digested and released by the addition of CaCl2. The fragmented chromatin was purified using the CUTANA DNA Purification Kit (EpiCypher 14-0050. Quantification, library preparation and sequencing was performed by the genomics core at DFCI.  DNA libraries were pooled and sequenced (2 x 150 bp paired end) on the Illumina NovaSeq 6000 platform.