Breeding mango is a long-term commitment, and from time of pollination to release of a new variety can take at least 20 years. Currently, in Australia, hand-pollination is used to target traits of known parents. Field assessment of hybrid progeny currently occurs without confirmation that the progeny are from the desired selected parents. Open pollination is a more cost-effective alternative to hand pollination; however, ensuring the desired parentage and, therefore, the inheritance of targeted traits is problematic. The use of molecular markers can confirm the parentage of the hybrids, with rogues eliminated at an early stage of the assessment. In this study, 12 microsatellite markers were used to determine parentage in hand- and open-pollinated mango hybrid progeny. Over 80% of hand-pollinated hybrid individuals were successfully identified for the most likely candidate father using a strict (95%) confidence level. In open-pollinated individuals, parentage was assigned with 95% confidence to 15 samples and to almost half the population with 80% confidence or more, assuming 75% of the candidate pollen donor population had been sampled. The result indicates that simple sequence repeat (SSR) analysis is an efficient method for cultivar identification. Although marker selection will not shorten the time to evaluate hybrid progeny, it can improve breeding efficiency by allowing the culling of unwanted hybrids prior to field evaluation. Development of additional molecular markers directly associated with traits of interest will allow further selection of hybrid populations for desirable traits prior to field planting.