ChIP was conducted using a ChIP assay kit (Cell Signaling Technology, USA) as described previously (Cui et al., 2018 (link)). For the KLF2 ChIP assay, DF1 cells were transfected with pCMV-Myc-KLF2 or an empty pCMV-Myc vector. The transfected cells were fixed in 1% formaldehyde for 10 min and quenched in 0.125 M glycine for 5 min. Then, 100–900 bp DNA/protein fragments were obtained through micrococcal nuclease digestion. The samples were immunoprecipitated with Myc-specific antibody (Cell Signaling Technology, USA) and mouse IgG (Beyotime, China) as a control to analyse the binding of KLF2 to the P1 promoter. The purified DNA was amplified by qPCR with the ChIP-qPCR primer shown in Table 1 . The input chromatin (2% nonimmunoprecipitated DNA) was used. The ChIP-qPCR data were calculated as previously detailed (Tatler et al., 2016 (link); Cui et al., 2018 (link)).
For the histone ChIP assay, ICP1 cells were transfected with pCMV-Myc-KLF2 or an empty pCMV-Myc vector. After 48 h of transfection, the cells were fixed and quenched as described above. The samples were immunoprecipitated with the H3K9me2 antibody (Abcam, UK) and the H3K27ac antibody (ABclonal, China), and mouse IgG (Beyotime, China) and rabbit IgG (Cell Signaling Technology, USA) were used as controls for the H3K9me2 and H3K27ac antibodies, respectively. The rest of the ChIP procedure was the same as described above.
For the histone ChIP assay, ICP1 cells were transfected with pCMV-Myc-KLF2 or an empty pCMV-Myc vector. After 48 h of transfection, the cells were fixed and quenched as described above. The samples were immunoprecipitated with the H3K9me2 antibody (Abcam, UK) and the H3K27ac antibody (ABclonal, China), and mouse IgG (Beyotime, China) and rabbit IgG (Cell Signaling Technology, USA) were used as controls for the H3K9me2 and H3K27ac antibodies, respectively. The rest of the ChIP procedure was the same as described above.