Anti trimethyl histone h3 lys4

ChIP assays were conducted as described previously (97 (link), 98 (link)). Cross-linked chromatin was immunoprecipitated with 5 μg of anti-Histone H3 (Abcam; catalog no.: ab1791), anti–trimethyl-Histone H3 Lys4 (Abcam; catalog no.: ab8580), anti–dimethyl-Histone H3 Lys9 (Abcam; catalog no.: ab1220), respectively, or negative control rabbit immunoglobulin G. Immunoprecipitated DNA was then purified and used as a template for PCR analysis. The primer sequences used for PCR are listed in Table S6.

Immunofluorescence was performed as described previously (99 (link)). Briefly, NIH3T3 cells were washed three times with cold PBS, fixed with 4% paraformaldehyde for 15 min, and permeabilized with 0.25% Triton X-100 (Sigma) for 10 min. Fixed cells were incubated with primary antibodies anti–trimethyl-Histone H3 Lys4 (Abcam; catalog no.: ab8580), anti–dimethyl-Histone H3 Lys9 (Abcam; catalog no.: ab1220), and Alexa Fluor 568–conjugated secondary antibodies step by step. Nuclei were stained with 4′,6-diamidino-2-phenylindole. Then the samples were imaged with fluorescence microscopy (Eclipse Ti2; Nikon) and processed using NIS-Element Version 5.02 software (Nikon, Melville, NY).