Gr antibody

ChIP material was prepared in accordance with the Magna ChIP (Millipore) manufacturer’s guidelines. Briefly, 1 × 10⁷ SKOV-3 cells were treated with 1 μM DEX or DMSO control for 3 h, and then ChIP assay was performed. IP was applied using 1 μg GR antibody (Cell Signaling) and equivalent amount of Rabbit IgG and H3 antibody (Millipore). PCR primers were designed on the promoter-3 region of ODZ4, and primer sequences used are provided in Supplementary Table 1. The PCR products were visualized using agarose gel electrophoresis or quantitative PCR. Experiments were repeated two times.

For Western blot studies, cells were collected, washed once with PBS, and then lysed in RIPA buffer (50 mM Tris-HCl, pH 8, 150 mM NaCl, 1% Triton X-100, 0.5% sodium deoxycholate, 0.1% SDS) with protease and phosphatase inhibitors (Thermo-Scientific, Waltham, MA, USA). Samples were boiled in Laemmli sample buffer for 5 min, and all samples were separated on an 8% or 10% SDS–polyacrylamide gel. Immunodetection of the respective proteins was performed using the antibodies described above. The anti-Flag monoclonal antibody (Sigma, F1804), β-catenin antibody (LifeSpan Biosciences; LS-C21245, Seattle, WA, USA), and GR antibody (Cell Signaling 3660S, Danvers, MA, USA) were used for these studies.