To understand the relationships among the genomes of Diospyros kaki (persimmon) and its wild relatives, we performed genomic in situ hybridization (GISH) on somatic metaphase chromosomes of D. kaki, using digoxigenin (DIG)- 11-UTP labeled DNA of eight Diospyros species. In a series of GISH, it appeared that the D. glandulosa probe hybridized to D. kaki chromosomes resulted with the strongest signal intensity. When comparable GISH was conducted, using unlabeled DNA of D. kaki as a blocking DNA, the highest concentration of blocking DNA was necessary to block hybridization of the D. glandulosa probe. These results suggest that the genomes of D. kaki and D. glandulosa share many common DNA sequences, and that the genome of D. kaki is most closely related to that of D. glandulosa among the species tested.