ChIP Experiments for C. elegans Proteins

ChIP experiments were performed as described previously with hypochlorite-isolated embryos or young adults (Guang et al. 2010 (link)). Animals were cross-linked in 2% formaldehyde for 30 min. Fixation was quenched with 0.125 M glycine for 5 min at room temperature. After cross-linking, samples were resuspended in FA buffer (50 mM Tris/HCl at pH 7.5, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, 150 mM NaCl) with a proteinase inhibitor tablet (Roche, 04693116001) and sonicated for 20 cycles at medium output (each cycle: 30 sec on and 30 sec off) with a Bioruptor 200. Lysates were precleared and then immunoprecipitated with 1.5 µL of anti-GFP antibody (Abcam, ab290) for SNPC-4, TOFU-4, and TOFU-5 and 5 µL of anti-PRDE-1 for PRDE-1 overnight at 4°C. Antibody-bound complexes were recovered with Dynabeads Protein A. Following extensive sequential washes with 150, 500, and 1 M NaCl, DNA was treated with RNase (Roche) and ProK (New England Biolabs). Finally, resulting DNA samples were purified with QIAquick PCR purification kit (Qiagen, 28104).