There is a growing appreciation for the importance of hybrid speciation in angiosperm evolution. Here, we show that Yucca gloriosa (Asparagaceae: Agavoideae) is the product of intersectional hybridization between Y. aloifolia and Y. filamentosa. These species, all named by Carl Linnaeus, exist in sympatry along the southeastern Atlantic coast of the United States. Yucca gloriosa was found to share a chloroplast haplotype with Y. aloifolia in all populations sampled. In contrast, nuclear gene-based microsatellite markers in Y. gloriosa are shared with both parents. The hybrid origin of Y. gloriosa is supported by multilocus analyses of the nuclear microsatellite markers including principal coordinates analysis (PCO), maximum-likelihood hybrid index scoring (HINDEX), and Bayesian cluster analysis (STRUCTURE). The putative parental species share only one allele at a single locus, suggesting there is little to no introgressive gene flow occurring between these species and Y. gloriosa. At the same time, diagnostic markers are segregating in Y. gloriosa populations. Lack of variation in the chloroplast of Y. aloifolia, the putative maternal parent, makes it difficult to rule out multiple hybrid origins of Y. gloriosa, but allelic variation at nuclear loci can be explained by a single hybrid origin of Y. gloriosa. Overall, these data provide strong support for the homoploid hybrid origin of Y. gloriosa.