Eight primer combinations that produced clear and a large number of polymorphic bands were screened from 64 EcoR I/Mse I primer combinations (Mse I fluorescent labeled). The genetic relationships of 21 ornamental cultivars of Ginkgo biloba L. from the United States of America, Holland, Japan, France, and China were analyzed. These primer combinations produced a total of 1 119 bands, 229 specific loci (including 54 absent bands, and 175 monomorphic bands). Among them, 983 polymorphic bands (PPB), accounting for 88%, were detected. The percentage of identification per primer combination was as high as 100%. The average PPB of 14 foreign cultivars was 35.86% and the average PPB of seven domestic cultivars was 31.51%. Genetic similarity coefficient (SC) among all cultivars varied from 0.4899 to 0.8499, and all cultivars were divided into the four clusters when SC was set at 0.7300. The cultivars from the same origin did not fall into the same group. The cultivars from France and China were classified into three groups. According to the comprehensive analyses based on specific loci, similarity coefficient, and clustering results, eight cultivars 'Fastigiata', 'Tit', 'Tubifolia', 'Daeryinxing', 'Variegata', 'Horizontalis, 'Pendula', and 'Yiyuanyeziyinxing' were considered to be important germplasms of ornamental cultivars of Ginkgo biloba.