Work

Protocol: Sonication-based Circular Chromosome Conformation Capture with next-generation sequencing analysis for the detection of chromatin interactions

Year: 2010

Type: article

Abstract: The three-dimensional organization of chromatin in the nuclear space is involved in regulation of gene expression.Circular Chromosome Conformation Capture \(4C) is an established method for genome-wide screening of chromatin interactions associated with a given locus of interest, without prior knowledge of the identity of interacting partners.Brie y, 4C involves the cross-linking of chromatin material, followed by restriction enzyme digestion of the chromatin, proximity-based ligation of interacting DNA fragments within the same DNA-protein complex, and ampli cation of interacting sequences by inverse PCR.The use of restriction enzyme digestion together with PCR could lead to biases in detection as well as false positives through repeated identi cation of clonal products.Here, we present a modi cation of the original 4C method, in which sonication is used to randomly fragment chromatin bers instead of restriction enzymes at speci c sites, to eliminate these biases, thus enabling high-throughput analysis by next-generation sequencing to detect interacting sequences.P.Y.H.H. and Y.H. made equal and critical

Source: Protocol Exchange

DOI: 10.1038/protex.2010.207