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PMID: 22903059 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Analysis of long-range chromatin interactions using Chromosome Conformation Capture.

Methods (San Diego, Calif.) ·Vol. 58 ·No. 3 ·2012-11-00 ·Pages 192-203

Naumova N, Smith EM, Zhan Y, Dekker J

Abstract

Chromosome Conformation Capture, or 3C, is a pioneering method for investigating the three-dimensional structure of chromatin. 3C is used to analyze long-range looping interactions between any pair of selected genomic loci. Most 3C studies focus on defined genomic regions of interest that can be up to several hundred Kb in size. The method has become widely adopted and has been modified to increase throughput to allow unbiased genome-wide analysis. These large-scale adaptations are presented in other articles in this issue of Methods. Here we describe the 3C procedure in detail, including the appropriate use of the technology, the experimental set-up, an optimized protocol and troubleshooting guide, and considerations for data analysis. The protocol described here contains previously unpublished improvements, which save time and reduce labor. We pay special attention to primer design, appropriate controls and data analysis. We include notes and discussion based on our extensive experience to help researchers understand the principles of 3C-based techniques and to avoid common pitfalls and mistakes. This paper represents a complete resource and detailed guide for anyone who desires to perform 3C.

MeSH Terms
Animals Cells, Cultured Chromatin/chemistry,genetics Chromosome Mapping/methods Cross-Linking Reagents/chemistry DNA Cleavage DNA Ligases/chemistry DNA Primers/genetics DNA Restriction Enzymes/chemistry Epistasis, Genetic Fixatives/chemistry Formaldehyde/chemistry Gene Library Humans Nucleic Acid Conformation Polymerase Chain Reaction
Chemicals
Chromatin Cross-Linking Reagents DNA Primers Fixatives Formaldehyde DNA Restriction Enzymes DNA Ligases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Naumova Natalia
Programs in Systems Biology and Gene Function and Expression, Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605-0103, USA.
Smith Emily M
Zhan Ye
Dekker Job
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Article Info
Journal
Methods (San Diego, Calif.)
Abbr.
Methods
ISSN
1095-9130
DOI
10.1016/j.ymeth.2012.07.022
Published
2012-11-00
Epub
2012-00-15
Pages
192-203
Language
English
Region
United States
NLM ID
9426302
PMCID
PMC3874837
Subset
IM
Grants
NHGRI NIH HHS · R01 HG003143 · United States
NHGRI NIH HHS · HG003143 · United States
NHGRI NIH HHS · HG004592 · United States
External Links
PubMed source DOI: 10.1016/j.ymeth.2012.07.022 PMC Full Text View journal details
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