Epitect chip antibody kit

ChIP assays were performed as described previously using human uveal melanoma C918 cell genomic DN.³⁰ (link) Rabbit polyclonal antiserum for Zeb1 as WB was used for immunoprecipitation whereas equal amount of pre-immune serum was used as a background control (IgG). The pan-histon 4 antibody (H4) from EpiTect ChIP antibody kit (Qiagen Cat. # GAH-2206) was used as a positive control. ChIP-PCR primer sequences and the expected PCR amplicon size are shown in Table S2.

ChIP assays using formaldehyde to cross-link genomic DNA were performed as we have described (37 (link), 55 (link)). The chromatin was mechanically sheared to an average length of 200 to 700 base pairs (bps). Rabbit polyclonal antiserum for Zeb1 was used for immunoprecipitation, whereas equal amount of preimmune serum was used as a control [immunoglobulin G (IgG)]. Immunoprecipitation with histone 3 antibody (H3) included in an EpiTect ChIP antibody kit (Qiagen, catalog no. GAH-2206) was used as a positive control. Sequences of primers for target promoters and the expected size of the PCR products are shown in table S1. ChIP-PCR programs were similar to that described below for qPCR, but with additional 1% bovine serum albumin and 1% dimethyl sulfoxide, and the PCR programs usually had a higher annealing temperature (e.g., 60° to 68°C) and longer extension time (e.g., 1 min). Results are representative of three independent experiments.