Figure 1.

Capturing spatial chromatin organization in vivo with 3C/5C technologies. (a) Current model of genome organization in the interphase nucleus. The diagram illustrates multiple levels of chromatin folding from the primary structural unit consisting of genomic DNA bound to nucleosomes (10 nm fiber; left). Secondary organization levels involve formation of 30 nm fibers through nucleosome-nucleosome interactions, and binding of individual fibers is believed to form tertiary structures (top). Folded chromatin occupies 'chromosome territories' represented by green, blue or orange shaded areas (right). Yellow circles indicate physical DNA contacts within (intra) or between (inter) chromosomes. (b) Schematic representation of 3C technology. 3C measures in vivo cross-linked DNA contacts at high resolution using individual PCR amplification and agarose gel detection. Interacting DNA segments located in cis is shown as an example to illustrate the 3C approach. Cis-interacting DNA fragments are represented by green and orange arrows and separated by a given genomic region (yellow line; left). Yellow circles represent cross-linked proteins. DNA segments are illustrated by arrows to highlight 'head-to-head' ligation configurations quantified by 3C. (c) Schematic representation of the 5C technology. 5C measures DNA contacts from 3C libraries using multiplex ligation-mediated amplification and microarray or high-throughput DNA sequencing. Genomic homology regions of 5C primers are shown in green and orange, and universal primer sequences are colored dark green or blue.