Abstract: Gobio acutipinnatus is a native and unique fish species in the Irtysh River Basin, as an important food source for carnivorous fish, it plays a crucial role in maintaining the biodiversity and ecological balance of the region. In order to understand the mitochondrial genome of G. acutipinnatus and clarify its phylogenetic position in the subfamily Gobioninae, we sequenced its complete mitochondrial genome and constructed a molecular phylogenetic tree of Gobioninae based on nucleotide sequences of 13 protein-coding genes. The results showed that the complete mitochondrial genome of G. acutipinnatus was 16608 bp in length, consisting 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, 1 non-coding region, and 1 origin of light-strand replication (OL) region. The base composition exhibits a significant AT preference (A+T=54.90%). Except for the protein-coding gene nad6 and 8 tRNA genes (trnQ, trnA, trnN, trnC, trnY, trnS2, trnE, and trnP) located outside the light chain, the remaining 28 genes were all located in the heavy chain. Among the 22 tRNA genes, only trnS1 gene lacked the DHU arm, while the other tRNAs could fold into the standard cloverleaf secondary structure, with a total of 57 G-U mismatches pairs. The phylogenetic tree (based on 13 protein-coding genes) revealed that the genus Saurogobio was embedded within Gobio, together with Mesogobio and Acanthogobio, they formed a branch. Gobio acutipinnatus clustered with Gobio occitaniae, Gobio cynocephalus, Gobio lozanoi, Gobio macrocephalus, Gobio soldatovi, and Gobio gobio within the genus Gobio. This study is the first to clarify the structural characteristics of the complete mitochondrial genome of G. acutipinnatus and determine its phylogenetic position, providing key molecular evidence for understanding the taxonomic relationships of this species within the genus Gobio and the Gobioninae subfamily.