GSM7631611: Inhibitor, DN3, ChIP, input; Mus musculus; ChIP-Seq
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody. Libraries were prepared using a Active Motive ChIP-IT ChIP-Seq kit (Cat# 53041). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, dNTP mix, DNA polymerase I Klenow and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3' end. Ligate adapter to DNA fragments in a cocktail containing DNA, quick ligation buffer, adapter oligo mix and T4 DNA ligase for 20 minutes at room temperature. After adapter ligation DNA was PCR amplified with PCR primers mix for 18 cycles and library fragments of 200-300 bp were band isolated from an agarose gel.