Abstract
The chromosome conformation capture (3C) technique is fundamental to many population-based methods investigating chromatin dynamics and organization in eukaryotes. Here, we provide a modified quantitative 3C (3C-qPCR) protocol for improved quantitative analyses of intra-chromosomal contacts. We also describe an algorithm for data normalization which allows more accurate comparisons between contact profiles.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Dekker J, Rippe K, Dekker M, Kleckner N (2002) Capturing chromosome conformation. Science 295:1306–1311
Dostie J, Richmond TA, Arnaout RA, Selzer RR, Lee WL, Honan TA, Rubio ED, Krumm A, Lamb J, Nusbaum C, Green RD, Dekker J (2006) Chromosome Conformation Capture Carbon Copy (5C): a massively parallel solution for mapping interactions between genomic elements. Genome Res 16:1299–1309
Simonis M, Klous P, Splinter E, Moshkin Y, Willemsen R, de Wit E, van Steensel B, de Laat W (2006) Nuclear organization of active and inactive chromatin domains uncovered by chromosome conformation capture-on-chip (4C). Nat Genet 38:1348–1354
Zhao Z, Tavoosidana G, Sjolinder M, Gondor A, Mariano P, Wang S, Kanduri C, Lezcano M, Sandhu KS, Singh U, Pant V, Tiwari V, Kurukuti S, Ohlsson R (2006) Circular chromosome conformation capture (4C) uncovers extensive networks of epigenetically regulated intra- and interchromosomal interactions. Nat Genet 38:1341–1347
Duan Z, Andronescu M, Schutz K, McIlwain S, Kim YJ, Lee C, Shendure J, Fields S, Blau CA, Noble WS (2010) A three-dimensional model of the yeast genome. Nature 465:363–367
Sexton T, Yaffe E, Kenigsberg E, Bantignies F, Leblanc B, Hoichman M, Parrinello H, Tanay A, Cavalli G (2012) Three-dimensional folding and functional organization principles of the Drosophila genome. Cell 148:458–472
Dixon JR, Selvaraj S, Yue F, Kim A, Li Y, Shen Y, Hu M, Liu JS, Ren B (2012) Topological domains in mammalian genomes identified by analysis of chromatin interactions. Nature 485:376–380
Lieberman-Aiden E, van Berkum NL, Williams L, Imakaev M, Ragoczy T, Telling A, Amit I, Lajoie BR, Sabo PJ, Dorschner MO, Sandstrom R, Bernstein B, Bender MA, Groudine M, Gnirke A, Stamatoyannopoulos J, Mirny LA, Lander ES, Dekker J (2009) Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science 326:289–293
Hagège H, Klous P, Braem C, Splinter E, Dekker J, Cathala G, de Laat W, Forné T (2007) Quantitative analysis of chromosome conformation capture assays (3C-qPCR). Nat Protoc 2:1722–1733
Court F, Miro J, Braem C, Lelay-Taha M-N, Brisebarre A, Atger F, Gostan T, Weber M, Cathala G, Forné T (2011) Modulated contact frequencies at gene-rich loci support a statistical helix model for mammalian chromatin organization. Genome Biol 12:R42
Court F, Baniol M, Hagège H, Petit JS, Lelay-Taha M-N, Carbonell F, Weber M, Cathala G, Forné T (2011) Long-range chromatin interactions at the mouse Igf2/H19 locus reveal a novel paternally expressed long non-coding RNA. Nucleic Acids Res 39:5893–5906
Braem C, Recolin B, Rancourt RC, Angiolini C, Barthes P, Branchu P, Court F, Cathala G, Ferguson-Smith AC, Forné T (2008) Genomic matrix attachment region and chromosome conformation capture quantitative real time PCR assays identify novel putative regulatory elements at the imprinted Dlk1/Gtl2 locus. J Biol Chem 283:18612–18620
Lutfalla G, Uzé G (2006) Performing quantitative reverse-transcribed polymerase chain reaction experiments. Methods Enzymol 410:386–400
Milligan L, Antoine E, Bisbal C, Weber M, Brunel C, Forné T, Cathala G (2000) H19 gene expression is up-regulated exclusively by stabilization of the RNA during muscle cell differentiation. Oncogene 19:5810–5816
Milligan L, Forné T, Antoine E, Weber M, Hemonnot B, Dandolo L, Brunel C, Cathala G (2002) Turnover of primary transcripts is a major step in the regulation of mouse H19 gene expression. EMBO Rep 3:774–779
Weber M, Hagège H, Lutfalla G, Dandolo L, Brunel C, Cathala G, Forné T (2003) A real-time polymerase chain reaction assay for quantification of allele ratios and correction of amplification bias. Anal Biochem 320:252–258
Acknowledgement
This work was supported by grants from the Institut National du Cancer [contract N° INCa_5960, PLBIO 2012-129, to T.F.], the Association pour la Recherche contre le Cancer [ARC contract n°SFI20101201555 to T.F.], the Ligue contre le cancer (comité Hérault), and the Centre National de la Recherche Scientifique (CNRS).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Ea, V., Court, F., Forné, T. (2015). Quantitative Analysis of Intra-chromosomal Contacts: The 3C-qPCR Method. In: Haggarty, P., Harrison, K. (eds) Population Epigenetics. Methods in Molecular Biology, vol 1589. Humana Press, New York, NY. https://doi.org/10.1007/7651_2015_269
Download citation
DOI: https://doi.org/10.1007/7651_2015_269
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6901-2
Online ISBN: 978-1-4939-6903-6
eBook Packages: Springer Protocols