title: Multiplexed chromosome conformation capture sequencing for rapid genome-scale high-resolution detection of long-range chromatin interactions.
abstract: Chromosome conformation capture (3C) technology is a powerful and increasingly popular tool for analyzing the spatial organization of genomes. Several 3C variants have been developed (e.g., 4C, 5C, ChIA-PET, Hi-C), allowing large-scale mapping of long-range genomic interactions. Here we describe multiplexed 3C sequencing (3C-seq), a 4C variant coupled to next-generation sequencing, allowing genome-scale detection of long-range interactions with candidate regions. Compared with several other available techniques, 3C-seq offers a superior resolution (typically single restriction fragment resolution; approximately 1-8 kb on average) and can be applied in a semi-high-throughput fashion. It allows the assessment of long-range interactions of up to 192 genes or regions of interest in parallel by multiplexing library sequencing. This renders multiplexed 3C-seq an inexpensive, quick (total hands-on time of 2 weeks) and efficient method that is ideal for the in-depth analysis of complex genetic loci. The preparation of multiplexed 3C-seq libraries can be performed by any investigator with basic skills in molecular biology techniques. Data analysis requires basic expertise in bioinformatics and in Linux and Python environments. The protocol describes all materials, critical steps and bioinformatics tools required for successful application of 3C-seq technology.
authors: Department of Cell Biology, Erasmus Medical Center, Rotterdam, The Netherlands.
authors: Ralph
authors: R
authors: Stadhouders
authors: Petros
authors: P
authors: Kolovos
authors: Rutger
authors: R
authors: Brouwer
authors: Jessica
authors: J
authors: Zuin
authors: Anita
authors: A
authors: van den Heuvel
authors: Christel
authors: C
authors: Kockx
authors: Robert-Jan
authors: RJ
authors: Palstra
authors: Kerstin S
authors: KS
authors: Wendt
authors: Frank
authors: F
authors: Grosveld
authors: Wilfred
authors: W
authors: van Ijcken
authors: Eric
authors: E
authors: Soler
mesh_terms: D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D002874:Chromosome Mapping / Q000379:methods*; D002875:Chromosomes / Q000737:chemistry / Q000378:metabolism; D019295:Computational Biology; D005557:Formaldehyde / Q000737:chemistry; D015698:Genomic Library; D023281:Genomics / Q000379:methods; D006801:Humans; D051379:Mice
chemical_list: D002843:Chromatin; D005557:Formaldehyde