Fish stocking, often from multiple source populations, is a common management practice frequently conducted without the means or effort to determine the reproductive contributions of stocked fish. Historically, the Minnesota Department of Natural Resources (MNDNR) has stocked four strains of muskellunge Esox masquinongy , but the contribution of these strains to current populations was unknown. Two strains came from Minnesota lakes, Shoepack Lake and Leech Lake, and the other strains came from Wisconsin and Iowa hatcheries and were of uncertain origin. The MNDNR discontinued stocking the Shoepack strain in the 1980s when that strain displayed poor growth in stocked waters. Managers were concerned that ancestry from this strain might be limiting the genetic potential for muskellunge to attain trophy size in stocked populations. Using 13 microsatellite DNA markers, we determined the ancestry of muskellunge in 10 supplemented native populations and 10 introduced populations. The ancestry from each of the four stocked strains of muskellunge was detected in some populations, but the level of ancestry was unrelated to the amount of stocking of a strain. Ancestry from native populations persisted in six of the supplemented populations despite years of stocking. The potential effects of Shoepack strain ancestry on fish size were limited in most lakes because of its low persistence. All stocked strains reproduced in at least some of the lakes, but some lakes had no evidence of reproduction by any stocked strain. Our results will help MNDNR manage genetic diversity among muskellunge populations and direct efforts toward appropriate actions to improve size structure. This study reinforces how genetic data are often useful for evaluating ancestry in stocked fish populations, whereas stocking histories may be poor indicators of current genetic composition.