GSM8001660: mTSC,H3K27ac,rin3_crispr,seq,2; Mus musculus; ChIP-Seq
For each ChIP experiment, 10 million TSCs were used. Cells were washed with PBS and cross-linked with 1% formaldehyde (Fisher Scientific) in PBS for 10 minutes at room temperature. The reaction was quenched with 125 mM glycine. Fixed cells were washed twice with cold PBS and resuspended in cold lysis buffer (15 mM HEPES pH 7.5, 140 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.5% N-lauroylsarcosine, 0.1% sodium deoxycholate and 0.1% SDS), incubated for 10 min on ice and sonicated with a Bioruptor Pico (Diagenode) for five cycles of 30s on and 30 s off.  The following antibodies were used: anti-Tfap2c (R & D systems, no. AF5059), anti-Tead4 (Abcam, no. ab58310), anti-Cdx2 (Bethyl Laboratories, no. A300-692A), anti-Gata3 (Cell Signaling, no. 5852T), anti-Yap1 (Cell Signaling, no. 14074S), anti-Pol II (Cell Signaling, no. D8L4Y), anti-H3K27ac (ChIP–seq) (Active motif, no. 39135). ChIP-nexus was performed according to He et al., 2015. ChIP-seq was performed as previously described in He et al., 2011, and included a whole cell extract (WCE). ChIP-nexus for transcription factors and ChIP-seq for histone modifications using standard Illumina protocols. ChIP-nexus reads were pre-processed by trimming off fixed and random barcodes and reassigning them to FASTQ read names.