Project description:Chromatin immunoprecipitation against CTCF followed by Illumina High-throughput sequencing. Examination of CTCF binding in red blood cells at 2 stages of development
Project description:Differential accessibility of DNA to nuclear proteins underlies the regulation of numerous cellular processes. DNA accessibility is primarily determined by the presence or absence of nucleosomes. At present, methods for mapping the positions and occupancy of nucleosomes genome-wide (MNase-seq) have uncovered the nucleosomal landscapes of different cell types and organisms. Conversely, methods specialized for the detection of large nucleosome-free regions of chromatin (DNase-seq, FAIRE-seq) have uncovered numerous gene regulatory elements. However, these methods are less successful in measuring the accessibility of DNA sequences within nucelosome arrays. Here we describe a new genome-wide method, RED-seq (deep sequencing analysis of Restriction Enzyme Digestion), that measures chromatin accessibility in an unbiased way. Using this method, we identified differences in chromatin accessibility between populations of cells, not only in broadly nucleosome-depleted regions of the genome (e.g., enhancers and promoters), but also within the more widespread regions of nucleosomal occupancy. Furthermore, RED-seq identified both large differences in chromatin accessibility in distinct cell lineages and subtle changes observed during differentiation of mouse embryonic stem cells (ESCs). Most notably, using RED-seq, we detected differences in accessibility among sequences occupied by distinct nucleosome variants. Therefore, RED-seq provides unique insights into genome-wide chromatin dynamics.
