The distribution of genetic variation within and among plant populations is influenced by both contemporary and historical factors. I used isozyme analysis of band phenotypes to examine genetic structure in the rare prairie forb Silene regia. Relationships between current-day population size, isolation, and phenotypic variation were assessed for 18 populations in two regions with differing postglacial history. Western populations from unglaciated southern Missouri and Arkansas were more genetically diverse based on the Shannon-Weaver index (H) and a polymorphic index than were more eastern populations. These differences may be due to loss of variation with repeated founding of new populations in previously glaciated sites in Indiana and Ohio. Within the western region, population size was not significantly correlated with genetic variation. In the east, size was correlated with Shannon-Weaver diversity. There was no relationship between variation and isolation in either region, but eastern populations were slightly more differentiated. Greater among-population differentiation and the demonstrated connection between population size and variation in the eastern sites may reflect lower levels of interpopulation gene flow in the fragmented remnant prairies of Indiana and Ohio.