H3k27me3 specific antibodies

We performed ChIP assays with the EZ‐ChIP KIT according to the manual (Millipore). EZH2, SUZ12 and H3K27me3‐specific antibodies were purchased from Millipore or mIgG/gIgG as negative control. Precipitated chromatin DNA was recovered and subjected to qRT‐PCR analysis. The primer sequences were enrolled Table S2.

Total histone protein pellet was dissolved in urea-CHAPS buffer (7 M urea, 2 M thiourea, and 4% CHAPS) and a mass of 10 μg of the protein was separated onto 15% SDS polyacrylamide gel. Subsequently, total histone protein was transferred onto a PVDF (Amersham, Little Chalfont, United Kingdom) membrane, which was processed for western blotting. The membrane was blocked in 5% skimmed milk for 2 h followed by three washings with PBST (1X PBS and 0.05% Tween20). The membrane was incubated in the primary antibody for 2–3 h. After the three washings, the membrane was incubated with anti-rabbit IgG secondary antibody labelled with horseradish peroxidase at a 1:10000 dilution (SIGMA, St. Louis, USA). The immunoblot was developed by applying the ECL method using Amersham ECL Prime Western Blotting Detection Reagent (Amersham, Little Chalfont, United Kingdom). For immunodetection, anti-H2B (ab1790; 1:5000 dilution), anti-H3-K4me3 (ab8580; 1:5000 dilution), anti-ubiquitinated H2B (MM0029; 1:500 dilution) and H3-K27me3 specific antibodies (02–449 Millipore, Massachusetts United State) were used.

Chromatin immunoprecipitation was performed with pluripotent ESCs and after differentiation in 3D-cultures according to a published protocol^{72 (link)}. Immunoprecipitation was conducted with polymerase 2 (Santa Cruz, Santa Cruz, USA; sc-899), H3K4me3 and H3K27me3 specific antibodies (Millipore, Germany; #17-614 and #17-622).
EpCAM-1 (ChIP): FW 5′-CGCAGCGCAAAGTCAAGTAT-3′; BW 5′- ACGTGAAAGCCGAAAGGGAT-3′
EpCAM-2 (ChIP): FW 5′- ATTGGAAAGTAAGCTGCCAGG-3′; BW 5′- GGACGAGACTGGACGTGAAA-3′
CenpI (ChIP): FW 5′- AACTCACGGATATTGAAGTGCAT-3′; BW 5′- CACAGAGCCAGGATACTGCTT-3′.