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Combining fluorescence imaging with Hi-C to study 3D genome architecture of the same single cell.

Accepted version
Peer-reviewed

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Type

Article

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Authors

Lando, David 
Riddell, Andy 
Wohlfahrt, Kai J 

Abstract

Fluorescence imaging and chromosome conformation capture assays such as Hi-C are key tools for studying genome organization. However, traditionally, they have been carried out independently, making integration of the two types of data difficult to perform. By trapping individual cell nuclei inside a well of a 384-well glass-bottom plate with an agarose pad, we have established a protocol that allows both fluorescence imaging and Hi-C processing to be carried out on the same single cell. The protocol identifies 30,000-100,000 chromosome contacts per single haploid genome in parallel with fluorescence images. Contacts can be used to calculate intact genome structures to better than 100-kb resolution, which can then be directly compared with the images. Preparation of 20 single-cell Hi-C libraries using this protocol takes 5 d of bench work by researchers experienced in molecular biology techniques. Image acquisition and analysis require basic understanding of fluorescence microscopy, and some bioinformatics knowledge is required to run the sequence-processing tools described here.

Description

Keywords

Animals, Cells, Cultured, Chromatin, Chromosomes, Imaging, Three-Dimensional, Mice, Molecular Biology, Molecular Conformation, Mouse Embryonic Stem Cells, Optical Imaging, Single-Cell Analysis

Journal Title

Nat Protoc

Conference Name

Journal ISSN

1754-2189
1750-2799

Volume Title

13

Publisher

Springer Science and Business Media LLC
Sponsorship
Medical Research Council (MR/M010082/1)
European Commission (277899)
Medical Research Council (MR/P019471/1)
Wellcome Trust (206291/Z/17/Z)
Wellcome Trust (098021/Z/11/Z)
Medical Research Council (MC_PC_12009)
Wellcome Trust (082010/Z/07/Z)