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The current Cucumis taxonomic classification places C. hystrix Chakr. in subgen. Cucumis based on its morphological similarities to cucumber (C. sativus L., 2n = 14). However, the chromosome number of C. hystrix was identified as 2n = 24, the same number as in subgen. Melo. Cucumis hystrix is therefore considered the first wild Cucumis species of Asiatic origin possessing 12 basic chromosomes. Thus, any research regarding its biosystematics would challenge the basic chromosome number and geographic location theories that govern the current taxonomic system. The production of the amphidiploid species (Cucumis ×hytivus Chen and Kirkbride, 2n = 38) obtained from the cross between C. hystrix and C. sativus and subsequent chromosome doubling would provide an effective means of investigating the relationship between Cucumis species with two different basic chromosome numbers. Thus, RAPD markers were used to study the taxonomic placement of C. hystrix and its interspecific hybrid with cucumber. Of the 220 arbitrary primers screened, 31 were used for analysis where 402 (96.3%) fragments were polymorphic among the germplasm examined. A UPGMA-based cluster analysis partitioned 31 accessions into two main groups [C. sativus (CS) and C. melo (CM)]. Under the similarity coefficient threshold of 0.23, these two groups can be further divided into five clusters with C. hystrix, C. ×hytivus, and C. sativus as separate clusters in the CS group. A modified taxonomic system is proposed based on these results and findings of a previous chloroplast DNA analysis with the genus Cucumis containing subgen. Cucumis with three species and subgen. Melo with six series.