RNA-Seq and ChIP-seq of HCC cells

For ChIP, approximately 2x107 cells were fixed using 1% formaldehyde before sonication. Coupled-DNA was eluted in 1% SDS/0.1M NaHCO3, and was reverse cross-linked at 65 oC, purified via phenol/chloroform extraction and ethanol precipitation, and finally subjected for semi- or realtime-qPCR. For Re-ChIP experiments, the complexes pulled down by the ChIP using the antibodies specific for the first protein were eluted by incubation 30 min at 37 oC in 10 mM DTT. After centrifugation, the supernatant was diluted 20 times with Re-ChIP buffer (1% Triton X-100, 2 mM EDTA, 150 mM NaCl, 20 mM Tris-HCl, pH 8.1), and subjected again into the ChIP procedure with the antibodies targeting against the second protein.For RNA-Seq, they were performed by Oebiotech.