Molecular markers are valuable tools in evaluating genetic diversity and fingerprinting plant germplasm. In this report, simple sequence repeat (SSR) markers were used for assessing genetic diversity in 41 dwarf and semidwarf and early flowering apple (Malus sp.) rootstocks. Sixty-two of 112 pairs of SSR primers generated multiple, scorable fragments. The total number of scored bands was 4138 with the polymorphic frequency ranging from 22.0% to 68.6% with a mean value of 58.5% in 737 alleles. The number of alleles per locus ranged from 6 to 19 with an average of 11.9 alleles. Polymorphic information content per locus was ranged from 0.176 to 0.885 with an average value of 0.606. These results suggested a complex genetic background and genetic diversity in these apple rootstocks. Based on three principal components and unweighted pair group mean average (UPGMA) of SSR data, the 41 apple rootstocks were divided into five groups. Group I contained M. xiaojinensis ‘Xiaojinhaitang'. Group II consisted of M. hupehensis var. pingyiensis ‘Pingyitiancha'. Group III contained M. baccata ‘Shandingzi' and its offspring. Group IV was composed of 16 apple rootstocks, including Malling and Malling Merton series from Great Britain; ‘Budagovski 9' from Russia; ‘Polish 22' from Poland; ‘Cornell-Geneva 24' from the United States; and ‘GM.256', ‘Nei Meng 11', ‘MD.001', ‘7734', and ‘7848' from China. Group V consisted of 16 Shao series rootstocks, which were offspring of M. honanensis × M. domestica ‘Ralls Genet'. This research suggests that the breeding can achieve best performance with more robust rootstock if crosses were performed among these five major groups of germplasms rather than within the major groups.