The invention relates to an identification method for seed plant species and application. The identification method comprises the following steps that the seed plant species are selected, ITS2 gene sequences of the seed plant species are obtained, an evolutionary tree is built based on the ITS2 gene sequences, the multiple pairs of sibling species are selected in the evolutionary tree, the genetic distance between each pair of sibling species is calculated, difference significance testing is conducted on the multiple groups of genetic distances of the species belonging to the same phylum, the genetic distances with non-significant differences are selected, the average value of the genetic distances with the non-significant differences is calculated, and thus the molecule critical value is obtained; and the genetic distance between the to-be-tested species and the target species is calculated, and the genetic distance and the molecule critical value of the phylum of the to-be-tested species are compared, so that the species category is determined. According to the identification method for the seed plant species, only ramification degree comparison needs to be conducted between to-be-tested samples or the suspected specifies and the ITS2 gene sequences of the target species, simpleness, convenience and easy operation are better achieved, accuracy is high, objectivity is high, the situation that the large number of other species in the same genus are sampled as a reference is not needed, and the high application value is achieved.