GSM8604796: Eu replaced with EF1a forward D345 cells, wapl, ChIP, rep2; Mus musculus; ChIP-Seq
Two million pro-B cells were initially crosslinked with 1.5 mM EGS at room temperature for 15 minutes with continuous rotation. This was followed by fixation with 1% formaldehyde for 15 minutes at room temperature, also with rotation, and quenched using 200 mM glycine. After washing twice with PBS, cells were then resuspended in a buffer containing 1% SDS and incubated for 10 min on ice before sonication. Chromatin was sonicated with Bioruptor (Diagenode, 30s on/ 30s off, 20 min on time) and diluted 10X with dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-HCl, PH8.0, 167 mM NaCl), and followed by immunoprecipitation with specific antibodies overnight at 4 °C The ChIP-seq libraries were prepared using the TruSeq Nano DNA Low Throughput Library Prep Kit (Illumina, Cat#20015964) according to the manufacturer's protocol. Samples were indexed using the TruSeq DNA Single Indexes Set A (Illumina, Cat#20015960) and pooled for sequencing.