Protein g agarose prepacked columns

Chromatin was prepared from hepatocytes for ChIP assays as previously described^{90 (link)}. Cells were fixed with 4% paraformaldehyde for 15 min, then glycine was added to a final concentration of 0.125 M and incubated for 10 min before harvesting. Chromatin was sonicated using a Bioruptor Pico (Diagenode, Denville, NJ, USA). Chromatin preparations were subjected to ChIP using a ChIP-IT High Sensitivity Kit and Protein G Agarose Prepacked Columns (Active Motif, Carlsbad, CA, USA) using either PPARα (Abcam Ab24509) antibody, normal rabbit IgG (Cell Signaling Technologies #2729S), or Histone H3 (Cell Signaling Technologies #4620) antibody. Rabbit IgG and Histone H3 were used as negative and positive controls, respectively. DNA was purified and concentrated using MinElute Reaction Cleanup columns (Qiagen). qRT-PCR and conventional PCR were performed using 2 μl of ChIP DNA samples from the 50 μl of purified samples using gene-specific primers (Supplementary Table 7). Cycle threshold (Ct) values of ChIP and input samples were calculated and presented as fold change.