Abstract: Mitochondrial 16S rRNA gene fragments were amplified from dolphins. Tursiops truncatus, Sousa chinensis and Steno bredanensis using PCR technique. The PCR products were about 600 bp and were directly sequenced subsequently. Finally, 515, 519 and 529 bp unambiguous nucleotide sequences were obtained respectively after excluding the primer sequences. The average base compositions were T 25.8%，C 20.7%，A 32.1% and G 21.4% with GC content being 42.1%. Alignment with 55 homologous sequences from 35 cetacean species in GenBank revealed 14 insertion/deletions and 127 variable sites (104 parsimony-informative sites and 23 singletons) among 497 truncated alignment sites. Sequence comparison suggested that Chinese T.truncatus and S. chinensis are different from the corresponding species abroad. NJ cluster analysis indicated that subclass Mysticeti is a monophyly whereas subclass Odontoceti is a polyphyly, including Delphinoidea, Physeteroidea (sperm whales), Ziphioidea and Platanistoidea. The sperm whales in Odontoceti was closely related with baleen whales in Mysticeti, and then clustered with Delphinoidea and then with Ziphioidea. The Platanistoidea was paraphyletic and located at the base of the tree, among which Platanistidae constitutes the base clade. Neophocaena phocaenoides(Phocoenidae) clustered with Delphinapterus leucas (Monodontidae) in Delphinoidea, inconsistent with morphological classification.