GSM9229104: Sox2 E14.5 ChIP-seq Input; Mus musculus; ChIP-Seq
Retinae dissected from C57/BL6 mice were flash-frozen and stored at −80°C for subsequent treatment. Retinae were thawed on ice and incubated with 1% formaldehyde in PBS at room temperature for 15 min. Fixation was stopped by the addition of 0.125 M glycine (final). Chromatin was isolated by the addition of lysis buffer and was sheared to an average length of 300-500 bp. Sheared chromatin was precleared with protein G agarose beads (Invitrogen). Chromatin immunoprecipitation (ChIP) was performed on 30-40 μg of sheared chromatin using 6 μl of SOX2 antibody (Active Motif # 39844). Complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment to collect DNA. Crosslinking was reversed by incubation overnight at 65°C, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.