Abstract: Fargesia rufa is the core staple bamboo species for giant pandas in the Minshan Mountains, whose population health and genetic characteristics directly underpin food supply and habitat stability for giant pandas. However, research on the genetic diversity of its natural populations remains relatively scarce. To elucidate the genetic diversity and population structure characteristics of natural F. rufa populations, this study employed SCoT molecular markers to conduct a genetic diversity analysis on 73 samples collected from 10 natural populations in the Giant Panda National Park of the Minshan Mountains. The results showed that a total of 23 effective primers with good polymorphism and high stability were screened from 112 SCoT primers, generating 249 clear DNA bands, of which 248 were polymorphic, with an average polymorphism ratio of 99.38%, indicating considerably high genetic diversity at the species level. Significant differences in genetic parameters were observed among populations. Populations QT4, QT2, and QT7 exhibited relatively higher genetic diversity, whereas population QT9 showed the most homogeneous genetic basis and markedly low diversity. The genetic distance among populations ranged from 0.0150 to 0.4923, with genetic similarity coefficients varying between 0.6112 and 0.9851. Cluster analysis classified all test materials into five major groups, revealing a certain correlation between population genetic relationships and geographic distribution patterns, along with evident genetic differentiation in some populations. The natural populations of F. rufa possessed abundant overall genetic variation, and SCoT markers were effective for genetic research on bamboo germplasm. These findings provided critical molecular genetic evidence for the conservation of F. rufa germplasm resources, population rejuvenation, ecological restoration and fine-scale management of giant panda habitats.