Combining fluorescence imaging with Hi-C to study 3D genome architecture of the same single cell.
Atkinson, Liam P
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Lando, D., Basu, S., Stevens, T. J., Riddell, A., Wohlfahrt, K., Cao, Y., Boucher, W., et al. (2018). Combining fluorescence imaging with Hi-C to study 3D genome architecture of the same single cell.. Nature protocols, 13 (5), 1034-1061. https://doi.org/10.1038/nprot.2018.017
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 the well of a 384 well glass bottomed 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 to 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 days of bench work by researchers experienced in molecular biology techniques. Image acquisition and analysis requires basic understanding of fluorescence microscopy, while some bioinformatics knowledge is required to run the sequence processing tools described here.
Cells, Cultured, Chromosomes, Chromatin, Animals, Mice, Imaging, Three-Dimensional, Molecular Biology, Molecular Conformation, Single-Cell Analysis, Optical Imaging, Mouse Embryonic Stem Cells
EC FP7 CP (277899)
Wellcome Trust (206291/Z/17/Z)
Wellcome Trust (098021/Z/11/Z)
Wellcome Trust (082010/Z/07/Z)
External DOI: https://doi.org/10.1038/nprot.2018.017
This record's URL: https://www.repository.cam.ac.uk/handle/1810/273627