Genetic characterization of germplasm is important for setting objective guidelines for conservation. One common problem found in genebanks is determining the value of populations with insufficient or unreliable data regarding their geographic origin. In this study, a genetic analysis based on RAPD markers was conducted to characterize a 'mystery' population of Solanum sucrense, a polysomic tetraploid potato (2n = 4x = 48), for which adequate documentation was lacking. The comparative analysis of genetic similarities between this mystery population and each one of 30 other S. sucrense populations in the genebank revealed that all populations within this species, including the mystery population, are significantly different from being duplicates, and are therefore worthy of separate conservation. RAPD markers also distinguished the mystery population from closely related tetraploid species S. oplocense, S. gourlayi and S. tuberosum ssp. andigena, suggesting that it is also not a duplicate of a population of these species. If RAPDs can clearly differentiate populations within highly heterogeneous tetraploids like S. sucrense, they should be generally useful for determining germplasm organization within potato species.