The Bagridae catfishes, belonging to the order Siluriformes and class Osteichthyes, are widely distributed in freshwater areas in Asia, such as China, Japan, Korean peninsula, and Vietnam. In China, Bagridae catfishes include more than 30 species across 4 genera. In recent years, due to environmental deterioration and human disturbance, populations of the native Bagridae had sharply decreased and some of them were difficult to collect. However, the classification of some Bagridae catfishes is ambiguous due to the difference of individuals, genders, or habitat. A species identification technique for various fishes based on DNA barcoding has been rapidly developed and widely employed. For many fishes, DNA barcoding requires additional validation before use. Herein, we have developed and evaluated DNA barcoding based on COI gene sequences for identifying Bagridae species. Altogether, 11 Bagridae catfish species were collected and sequenced with the standard DNA barcoding protocols, then combined with 8 available species acquired from the GenBank. A total of 60 sequences from 19 species pertaining to 4 genera were analyzed. DnaSP 5 and MEGA7.0 were employed to calculate the base composition, sequence characterization, and genetic distance, and construct the neighbor-joining phylogenetic tree. The mrbayes software was used to construct the Bayesian tree. The result showed that the mean base contents were:24.82% A, 30.44% T, 27.10% C, and 17.64% T. AT contents (55.26) were higher and the base composition was biased as others teleost fishes were also commonly observed. The nucleotide sequences included 226 variant sites, 195 parsimony informative sites, and 31 singleton sites. The average transition/transversion ratio was 3.35. Genetic distances within species, genus, and families were 0.0041, 0.1136, and 0.1268, respectively. The ratio of genetic distances between interspecies and intraspecies was 27.7, which indicated a notable genetic difference of the COI gene sequence. The efficiency of DNA barcoding based on the COI gene was reaffirmed from its successful identification of all 19 Bagridae catfishes in this study. This classification result is consistent with their morphologies. However, only catfishes can be clustered into a single phylogenetic tree branch. Other Bagridae species were intertwined, and their phylogenetic status needs further research combined with data from more samples. The results revealed that the COI gene was suitable as a DNA barcoding parameter for identifying species; it provided a useful reference for the phylogenetic relationships of Bagridae.