This study used the sequences of two mitochondrial genes, ATPase 6 and ATPase 8, to analyze the genetic diversity and population structure of Cirrhinus mrigala from four geographically distinct river basins in Pakistan's Indus River System. Following the collection of 20 samples from the Taunsa, Chenab, Jhelum, and Ravi rivers, DNA was extracted, amplified by PCR, and sequenced. Genetic heterogeneity was observed among individuals and populations, as sequence alignment revealed a few nucleotide variations in the ATPase 6 and ATPase 8 genes. Phylogenetic analyses showed that the Taunsa and Chenab samples appeared to form a population-specific group. However, the genetic relationships among the populations were very close, suggesting a common origin. High genetic similarity and gene flow across geographically dispersed groups were demonstrated by low genetic distances between many samples. In summary, the findings indicate that C. mrigala has comparatively little population differentiation but substantial genetic variation within populations. This information can be useful for comprehending population connectivity, genetic conservation, fisheries management, and the sustainable use of this economically significant fish species in Pakistan.