In this study, a fingerprinting system for 16 Michelia germplasms was established using high-throughput genotyping of fluorescent simple sequence repeat (SSR) markers. In total, 102 alleles were detected using 13 pairs of SSR markers from closely related species, and the number of alleles per locus ranged from 5 to 11, with an average of 7.8 alleles. Observed heterozygosity (Ho), expected heterozygosity (He), and polymorphic information content (PIC) ranged from 0.1250 to 0.5625 (mean 0.3650), 0.6703 to 0.9113 (mean 0.8099), and 0.5748 to 0.8714 (mean 0.7515), respectively. Among the four selected core primer pairs, the combinations LT106 and SGA5, LT106 and LT58, and SGA5 and MMA51 could unambiguously distinguish 16 Michelia germplasms. A cluster analysis showed the similarity coefficient of the 16 Michelia germplasms to range from 0.70 to 0.90, and different individuals from the same species clustered in the same branch. The fluorescence SSR genotyping system established in this study was efficient, rapid, and accurate; moreover, this approach provides a theoretical basis for identifying germplasms and for the protection of new varieties of the genus Michelia , and it provides a robust foundation for further breeding development of Michelia sp.