The three-dimensional (3D) organization of genomes refers to the spatial organization and folding of chromatin within the nucleus of cells. Over the years, the initial development and subsequent applications of chromosome capture techniques (3C) and their derivatives have permitted the study of the 3D genome organization at the deepest level of resolution. In this chapter, we provide a practical guide for the analysis and interpretation of Hi-C data, along with applications for detecting chromosomal reorganizations, such as Robertsonian fusions.