In vertebrates, two condensin complexes exist, condensin I and II, which have differing but unresolved roles in organizing mitotic chromosomes. To accurately dissect the role of each complex in mitosis, we have made and studied the first vertebrate conditional knockouts of the condensin I subunit CAP-H and condensin II subunit CAP-D3 in chicken DT40 cells. Live-cell imaging reveals highly distinct segregation defects. CAP-D3/condensin II KO results in masses of chromatin forming anaphase bridges. CAP-H/condensin I KO anaphases have a more subtle defect, with chromatids showing fine chromatin fibres that are associated with failure of cytokinesis and cell death. Super-resolution microscopy reveals condensin I-depleted mitotic chromosomes as wider and shorter with a diffuse chromosome scaffold, while condensin II-depleted chromosomes retain a more defined scaffold with chromosomes more stretched and seemingly lacking in axial rigidity. We conclude condensin II is required primarily to provide rigidity by establishing an initial chromosome axis around which condensin I can arrange loops of chromatin.