This study combined population genetics and parentage analysis to obtain foundational data for the conservation of Magnolia kwangsiensis. M. kwangsiensis is a Class I tree species that occurs in two disjunct regions in a biodiversity hotspot in southwest China. We assessed the genetic diversity and structure of this species across its distribution range to support its conservation management. Genetic diversity and population structure of 529 individuals sampled from 14 populations were investigated using seven nuclear simple sequence repeat (nSSR) markers and three chloroplast DNA (cpDNA) fragments. Parentage analysis was used to evaluate the pollen and seed dispersal distances. The nSSR marker analysis revealed a high genetic diversity in M. kwangsiensis, with an average observed (Ho) and expected heterozygosities (He) of 0.726 and 0.687, respectively. The mean and maximum pollen and seed dispersal distances were 66.4 and 95.7 m and 535.4 and 553.8 m, respectively. Our data revealed two distinct genetic groups, consistent with the disjunct geographical distribution of the M. kwangsiensis populations. Both pollen and seed dispersal movements help maintain genetic connectivity among M. kwangsiensis populations, contributing to high levels of genetic diversity. Both genetically differentiated groups corresponding to the two disjunct regions should be recognized as separate conservation units.