Mapping networks of physical interactions between genomic elements using 5C technology
Program in Gene Function and Expression
Chromatin; Gene Library; Genomics; Molecular Conformation; Polymerase Chain Reaction
Genetics and Genomics
Genomic elements separated by large genomic distances can physically interact to mediate long-range gene regulation and other chromosomal processes. Interactions between genomic elements can be detected using the chromosome conformation capture (3C) technology. We recently developed a high-throughput adaptation of 3C, 3C-carbon copy (5C), that is used to measure networks of millions of chromatin interactions in parallel. As in 3C, cells are treated with formaldehyde to cross-link chromatin interactions. The chromatin is solubilized, digested with a restriction enzyme and ligated at low DNA concentration to promote intra-molecular ligation of cross-linked DNA fragments. Ligation products are subsequently purified to generate a 3C library. The 5C technology then employs highly multiplexed ligation-mediated amplification (LMA) to detect and amplify 3C ligation junctions. The resulting 5C library of ligated primers is analyzed using either microarray detection or ultra-high-throughput DNA sequencing. The 5C protocol described here can be completed in 13 d.
DOI of Published Version
Nat Protoc. 2007;2(4):988-1002. Link to article on publisher's site
Dostie, Josee and Dekker, Job, "Mapping networks of physical interactions between genomic elements using 5C technology" (2007). Program in Gene Function and Expression Publications and Presentations. 107.