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 J, Dekker J. (2007). Mapping networks of physical interactions between genomic elements using 5C technology. Program in Gene Function and Expression Publications. https://doi.org/10.1038/nprot.2007.116. Retrieved from https://escholarship.umassmed.edu/pgfe_pp/107