ChIP-Atlas: SRX4318734

Antigen

library_strategy: ChIP-Seq
library_source: GENOMIC
library_selection: ChIP
library_construction_protocol: The normal cells were dissected and FACS isolated from primary mammary tissue. The PY230 cells were transplanted into syngeneic mice to form tumors, and the tumors were dissected and FACS isolated. See the Methods section of the manuscript for details of isolation and FACS markers. For H3K27ac ChIP, around 20k cells were crosslinked with 1% PFA at room temperature for 10 minutes, sonicated with Covaris M220 into 200-700 bp fragments, and incubated with anti-H3K27ac antibody (Diagenode C15410174) overnight at 4¡C. Protein A beads pull down (Invitrogen 10001D), washing, on-beads tagmentation (Illumina Nextera FC-121-1030), reverse crosslinking, library amplification (13 PCR cycles) and DNA purification were performed as described. The input library was prepared by tagmentation of 1 ng of reverse crosslinked input DNA and then amplified and processed together with the ChIP DNA. Multiple input preparations were pooled together to ensure sufficient complexity of the input library. For the SOX10 ChIP, 500k control and SOX10-AVI/BirA cells were processed as above, except that streptavidin beads (Pierce #88816) were used for pull down, and that two additional washing steps with 2% SDS in PBS were performed to significantly remove ChIP backgrounds. The input library was prepared as above. ChIPmentation method (Schmidl et al., 2015)