What is the 3C Method?
What is the 3C Method?
Chromatin Conformation Capture (3C) is an important technique used to study chromatin structure, as well as the basis for several other derivative techniques. The protocol involves formaldehyde cross-linking of cells followed by chromatin isolation and digestion with a restriction enzyme.
What is Hi-C sequencing?
Hi-C uses high-throughput sequencing to find the nucleotide sequence of fragments and uses paired end sequencing, which retrieves a short sequence from each end of each ligated fragment. The pair of sequences are individually aligned to the genome, thus determining the fragments involved in that ligation event.
What is 4C sequencing?
4C-seq is a derivative 3C method, designed to search the genome for sequences contacting a selected genomic site of interest. 4C-seq employs inverse PCR and next generation sequencing to amplify, identify and quantify its proximity ligated DNA fragments.
What is Hi-C genetics?
In Hi-C, cells are fixed with formaldehyde, causing interacting loci to be bound to one another by means of covalent DNA-protein cross-links. When the DNA is subsequently fragmented with a restriction enzyme, these loci remain linked. A biotinylated residue is incorporated as the 5′ overhangs are filled in.
How is capture C used in 3C seq?
Capture-C adds an additional pull-down of the biotinylated fragments with magnetic beads to the 3C method. A new refinement of the Capture-C method (NG Capture-C) is available. The Hi-C approach extends 3C-Seq to map chromatin contacts genome-wide, and it has also been applied to studying in situ chromatin interactions.
How are 3C and 3C based techniques used?
Recently, chromosome conformation capture (3C) and 3C-based techniques using high-throughput sequencing data have emerged as powerful tools to reconstruct the spatial topology at regional, whole chromosome and genome levels [ 40, 41, 42, 43, 44, 45 ].
How are 3C and 4C technologies used in DNA sequencing?
4C technology was developed by combining 3C with microarray [ 60, 61] or, more recently, next-generation sequencing (NGS) technologies [ 62, 63 ]. This method is able to assess chromatin interactions between one genomic locus of interest (referred to as bait or viewpoint) and all other genomic loci (one versus all) (Fig. 2b ).
What’s the difference between Hi-C and 3C seq?
Hi-C, 3C-Seq, and Capture-C comprise a family of methods for analyzing chromatin interactions. Capture-C adds an additional pull-down of the biotinylated fragments with magnetic beads to the 3C method.