GSM5620516: Cauda sperm DNA - H3K27ac_rep1; Mus musculus; ChIP-Seq
Sperm histone ChIP was carried out using a previously published protocol (Hisano et al., 2013). Briefly, for each individual biological rep, sperm collected from the Caputs and Cauda of 10-15 mice were pooled, purified and pre-treated with 50μM DTT for 2h at RT to open sperm chromatin. Cell suspensions were split into roughly 15-20 million sperm aliquots per IP and treated with 90 U of MNase (New England Biolabs, #M0247S) for 5 min at 37C. MNase-digested chromatin was then pre-cleared with blocked A/G Sepharose beads for 1h at 4C. For IP, 5 μg of antibody (H3K27ac or H3K9me2) was added to the pre-cleared chromatin overnight followed by 50 μL of blocked beads for 4 h at 4C. Beads were washed and DNA was then eluted in elution buffer. Following RNase A and proteinase K treatment, DNA was purified using phenol/chloroform isolation and ethanol precipitation and resuspended in 40 μL ultrapure water. Library preparation using 10ng of DNA and 75 bp paired-end sequencing with at least 50 million reads per samples was carried out at the UT Health Cancer Genomics Center, Houston.
GEO Accession
GSM5620516