Rana kukunoris is an amphibian species endemic to the Qinghai-Xizang Plateau and adapted to high-elevation environments. To clarify its genetic diversity and phylogeographic structure, mitochondrial cytochrome b (Cytb) sequences from multiple localities across the northeastern plateau were analyzed. The results indicate moderate haplotype diversity coupled with low nucleotide variation, suggesting a demographic history shaped by historical bottlenecks and limited expansion. Furthermore, population genetic analyses reveal restricted gene flow and notable genetic differentiation among geographically separated localities. Phylogenetic reconstruction supports the presence of two major lineages, corresponding to northern and southern groups divided by an elevational boundary at approximately 3200 m. Historical demographic inference further suggests regionally distinct population dynamics rather than a pronounced, synchronous expansion. Overall, these findings demonstrate that plateau topography and Quaternary climatic oscillations have played key roles in shaping the evolutionary history and current genetic structure of R. kukunoris.