Single-Nucleosome Mapping of Histone Modifications in S. cerevisiae
Figure 1
(A) Nucleosomes are first cross-linked to DNA using formaldehyde. Cross-linked chromatin is digested to mononucleosomes with micrococcal nuclease. Mononucleosomal digests are immunoprecipitated using an antibody specific to a particular histone modification, and immunoprecipitated DNA is isolated and labelled with Cy5. DNA is also isolated from the same nuclease titration step prior to immunoprecipitation, labelled with Cy3, and mixed with Cy5-labeled immunoprecipitated DNA. Labelled DNA is then hybridized to a tiled microarray covering half a megabase of yeast genome.
(B) Example of raw data. Data are shown for all modifications tested, along with PolII data. Red (green) indicates enrichment (depletion), while grey indicates missing data. Data from probes found in linker regions are not shown. Each row represents median data from multiple replicates with one antibody, as indicated (PanAc refers to a nonspecific antibody to acetyl-lysine, which we used to measure bulk acetylation). “Nucleosomes” shows positions of nucleosomes previously described [29], with dark brown for well-positioned nucleosomes, very light brown for linkers, and intermediate brown for delocalized nucleosomes. “ORFs” shows locations of annotated genes. Data shown are for Chromosome III coordinates 58,900 to 72,100.