The invention discloses a method for breeding zebra fish with wnt16 gene deletion through gene knockout. Compared with the prior art, the method has the advantages that specific genes can be more efficiently and more accurately silenced in a genome of a living body, in addition, the preparation is simple, the cost is low, moreover, a plurality of sites on a target gene can be simultaneously shorn, and single genes of any number can be silenced. The method is used for carrying out the relevant researches on gene and skeletal development, also can be used for carrying out the exploration research of other aspects, is used for detecting that whether the deletion of the wnt16 gene has relevance with the development of other organs, such as the heart, or not, and has a very good medical research value, meanwhile, the zebra fish with the wnt16 gene knocked out has the obviously shortened growth cycle, and thus the commercial value is good.