GSM1446977: ESCs_Pol II; Mus musculus; ChIP-Seq
Harvested cells were lysated and sonicated to 100-500bp. Protein-DNA complexes were isolated with antibody. ChIP complexes were eluted with 1% SDS in 100 mM NaHCO3 and crosslinks were reversed by incubating samples over night at 65C. DNA fragments were purified using phenol-chloroform extraction and ethanol precipitation Libraries were constructed according to the Illumina ChIP-seq library preparation instructions. Briefly, pull-down DNA was end repaired using a combination of T4 DNA polymerase, T4 PNK and Klenow polymerase. The DNA was then purified and used for A-tailing by adding a dATP to the 3' end. An indexed adaptor with a "T" overhang at the 3' end was then ligated to each DNA sample. The ligated DNA was then subjected to PCR amplification followed by gel purification.