Anti jarid2

Immunoblotting was performed as previously described [22 (link)]. Anti-Flag (Cat. F7425) was purchased from Sigma. Anti-GAPDH (Cat. sc-47724) was purchased from Santa Cruz Biotech. Anti-Jarid2 (Cat. ab48137) and anti-p16 (Cat. ab201980) were purchased from Abcam.

Cells were directly lysed and subjected to 10% SDS-PAGE. Immunoblotting was performed after transferring the proteins onto nitrocellulose membranes (Schleicher & Schuell Microscience) with a Mini Trans-Blot apparatus (Bio-Rad). After 2 h of blocking, the membranes were incubated overnight at 4°C with the following specific primary antibodies: anti-Jarid2 (Abcam), anti-p-STAT3, anti-STAT3 (Cell Signaling), and anti-β-actin Abs (Sigma-Aldrich). After the membranes were washed, subsequent incubations with the appropriate HRP-conjugated secondary antibodies were conducted for 1 h at room temperature; after extensive washing, the signals were visualized with an ECL substrate (Pierce Chemical).

ChIP experiments were performed as previously described [19 (link)]. The crosslinked chromatins were immunopreciptated with anti-H3K27me3 (Cat. ab6002) and anti-Jarid2 (Cat. ab48137) antibodies from Abcam. The enrichment of the specific amplified region was analyzed by quantitative PCR and percentage enrichment of each modification over input chromatin DNA was shown. ChIP PCR primers sequences are listed in the Supplementary Materials.