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Background: Many endangered species exist in small, genetically depauperate, or inbred populations, hence promoting genetic differentiation and reducing long-term population viability. Forest Musk Deer (Moschus berezovskii) has been subject to illegal hunting for hundreds of years due to the medical and commercial values of musk, resulting in a significant decline in population size. However, it is still unclear to what extent the genetic exchange and inbreeding levels are between geographically isolated populations. By using whole-genome data, we reconstructed the demographic history, evaluated genetic diversity, and characterized the population genetic structure of Forest Musk Deer from one wild population in Sichuan Province and two captive populations from two ex-situ centers in Shaanxi Province. Results: SNP calling by GATK resulted in a total of 44,008,662 SNPs. Principal component analysis (PCA), phylogenetic tree (NJ tree), ancestral component analysis (ADMIXTURE) and the ABBA-BABA test separated Sichuan and Shaanxi Forest Musk Deer as two genetic clusters, but no obvious genetic differentiation was observed between the two captive populations. The average pairwise FST value between the populations in Sichuan and Shaanxi ranged from 0.05–0.07, suggesting a low to moderate genetic differentiation. The mean heterozygous SNPs rate was 0.14% (0.11%—0.15%) for Forest Musk Deer at the genomic scale, and varied significantly among three populations (Chi-square = 1.22, p < 0.05, Kruskal–Wallis Test), with the Sichuan population having the lowest (0.11%). The nucleotide diversity of three populations varied significantly (p < 0.05, Kruskal–Wallis Test), with the Sichuan population having the lowest genetic θπ (1.69 × 10–3). Conclusions: Genetic diversity of Forest Musk Deer was moderate at the genomic scale compared with other endangered species. Genetic differentiation between populations in Sichuan and Shaanxi may not only result from historical biogeographical factors but also be associated with contemporary human disturbances. Our findings provide scientific aid for the conservation and management of Forest Musk Deer. They can extend the proposed measures at the genomic level to apply to other musk deer species worldwide. [ABSTRACT FROM AUTHOR]