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Understanding the genetic variability within a species population is important for elucidating adaptive divergence and potential mechanisms involved in speciation. Excluding geographical limitations, historical events and human disturbance, complex environmental conditions also influence the amount of gene flow among populations. We investigated the relationship between the genetic variability of roe deer (Capreolus pygargus) and environmental factors using molecular techniques and mathematical statistical methods. We compared 460 base pairs from the mitochondrial DNA (mtDNA) control region of 88 faecal samples individuals across eight different geographic regions in Northeastern China. Concurrently, we correlated seven environmental factors: latitude, longitude, altitude, temperature, precipitation, frost-free period and vegetation types, with genetic variability within roe deer populations. Our study suggests that climatic conditions and genetic diversity of roe deer were more closely correlated than geographic location and habitat type, with a cumulative variance of 43.6%. From the point of view of climate change to analysis, climatically stable regions are expected to favor diversification processes and sustain genetically more diverse populations by comparison with areas that have experienced changeable climatic conditions. Simultaneously, we found that roe deer dispersal was associated with periods of reproduction in vegetation type influenced by seasonal change. So, we suggest that precipitation is the major factor that indirectly affects the genetic difference within roe deer populations.