The alterations of hierarchical structures of bone by gene mutation in the zebrafish, which is associated with abnormal bone mineralization and bone disease, were reported for the first time in this paper. Bone samples from the liliput(dtc232) (lil) mutants as well as normal controls were studied by polarized light microscope, scanning electron microscope (SEM), transmission electron microscope (TEM), and atomic force microscope (AFM). Light microscopy examinations reveal that the lil bone has asymmetric mineralization and much thinner bone wall. The SEM studies show a lot of microcracks in lil bone wall. And the plywood-like structure of the normal bone does not exist in the lil bone, which is confirmed by the measurements of polarized light microscope. Furthermore, the TEM investigations display the collagen fibrils with two typical diameters. For the thinner collagen fibrils, the diameter of lil bone is about twice larger than that of the wild-type bone. And for the thicker one, there is a small increase in diameter after mutation and the band periodicity of the lil bone is similar with that of wild-type bone, which is consistent with the result of AFM. The morphologies of the minerals revealed that the mutated mineral was in bigger size and the shape was irregular but not plate-shaped.