Fishes of the family Muraenidae, commonly known as morays, are of commercial importance to fisheries worldwide. Morphologically, classification and identification in Muraenidae fish remain problematic because many closely-related species share common overlapping color patterns. Additionally, colorations and patterns in some Muraenidae species change greatly during their lives, making the juveniles quite distinct from their adult forms. To analyze the phylogenetic relationships of the Muraenidae fish based on the molecular level, and to clarify the species taxonomic controversies, 16S rRNA sequences of 26 Muraenidae species of 9 genera from China Sea were obtained by PCR amplification in this study. Combined with other Muraenidae sequences obtained from GenBank, the base composition, conserved sites, variable sites and genetic distance of the sequences were analyzed using Mega 7.0 software and the molecular phylogenetic trees were constructed by maximum likelihood and Bayesian inference methods. Results showed that: In the genetic distance results, the genetic distance values of 35 Muraenidae species ranged from 0.009 to 0.309. The maximum value (0.309) was found between species Gymnothorax pesudothyrsoideus and Scuticaria tigrina, while the minimum value (0.009) was between G. kidako and G. niphostigmus. Within the genetic distance among 10 genera，the maximum inter-genera value (0.187-0.279) was found between genera Gymnothorax and Uropterygius, while the minimum value (0.045-0.131) was between genera Enchelycore and Muraena. Additionally, certain inter-species distance values within the same genus were large and even exceeded the inter-genera values. Like some species in the genera Gymnothorax, Echidna and Enchelycore, it indicated that the genetic differentiation level in some species had exceeded the inter-genera levels. In the phylogenetic tree, Muraenidae species formed two major groups: subfamily Uropterygiinae and subfamily Muraeninae groups. Uropterygiinae group comprised two genera Uropterygius and Scuticaria, while Muraeninae group comprised 8 genera including Gymnothorax, Echidna, Enchelycore, Gymnomuraena, etc., which was consistent with the current morphological classification result. In Uropterygiinae group, two genera Uropterygius and Scuticaria formed a parallel sister branch. In Muraeninae group, species G. zebra formed a separate branch and was located at the base of the group, and the remaining species were clustered into another large branch. Within this branch, genera like Gymnothorax, Echidna, Enchelycore and Muraena could not form monophyletic group. Species in these genera were clustered together in different positions of the phylogenetic tree, revealing their polyphyletic evolutionary status, which was consistent with the recent molecular phylogenetic studies. For the species/genera that were in morphological classificational controversy, our phylogenetic results supported the view that G. zebra was classified into genus Gymnomuraena as a monotypic species. Scuticaria tigrina was classified into genus Scuticaria and its phylogenetic status was sister to the genus Uropterygius. Species Pseudechidna brummeri showed distant relationship with genus Strophidon, which might be separated from Strophidon and classified into genus Pseudechidna. Enchelycore pardalis was closely related with genus Muraena at the molecular level, but morphologically, certain Enchelycore basic characteristics also existed in the species. Whether it could be classified into genus Muraena, further comprehensive molecular and morphological studies are needed for confirmation.