GSM2771238: Prdm16 ChIP-Seq; Mus musculus; ChIP-Seq
BAT from 10–15 mice (male, 8–10 wk old) were dissected and homogenized with a douncer. Paraformaldehyde was added to the homogenate to 1% final concentration. Homogenization was performed prior to cross-linking because it is difficult to collect mature brown adipocytes by centrifugation. The homogenate was then quenched with 125 mM glycine, and the nuclei were collected  by centrifugation. The nuclei were lysed with lysis buffer (1% SDS, 50 mM Tris-HCl, 10 mM EDTA at pH 8.0) and sonicated to yield an average size of DNA fragment of 150–200 base pairs (bp). The lysate was diluted 10-fold with a dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-HCl, 167 mM NaCl at pH 8.0) and incubated with the desired antibody overnight. The protein–DNA complex was isolated with protein G Dynabeads, washed, and eluted off the beads. The eluent was treated with RNase A and proteinase K before reversing crosslinking.DNA was then harvested by ethanol precipitation. The library was prepared with the NEBNext ChIP-seq library preparation master mix set for Illumina (New England Biolabs, E6240S).
GEO Accession
GSM2771238