The process of cellular differentiation requires the distinct spatial organization of the microtubule cytoskeleton, the arrangement of which is specific to cell type. Microtubule patterning does not occur randomly, but is imparted by distinct subcellular sites called microtubule-organizing centers (MTOCs). Since the discovery of MTOCs fifty years ago, their study has largely focused on the centrosome. All animal cells use centrosomes as MTOCs during mitosis. However in many differentiated cells, MTOC function is reassigned to non-centrosomal sites to generate non-radial microtubule organization better suited for new cell functions, such as mechanical support or intracellular transport. Here, we review the current understanding of non-centrosomal MTOCs (ncMTOCs) and the mechanisms by which they form in differentiating animal cells.