Inter-simple sequence repeat (ISSR) amplification was evaluates for its usefulness in generating DNA markers for sweetpotato and related wid species. ISSR markers were obtained through PCR amplification using simple sequence repeat (SSR) primers. Optimization of the reaction conditions was successfully achieved fro 24% out of 100 SSR primers screened. The functional primers included anchored and unanchored primers. All of the anchored primers used were dinucleotide repeat motifs. The unanchored primers consisted of tri-, tetra- and penta-nucleotide repeat motifs. Eight primers were selected and employed to assess the genetic diversity and relationship between 34 accessions of sweetpotato and its related wild species. The ISSR markers are highly polymorphic among the ta a studied. Several sweetpotato accessions were clustered together based on their geographic origin. The mode of inheritance of the ISSR markers was analyzed in a pseudo-test cross mapping population. Twenty-two SSR primers amplified 70 reproducible polymorphic ISSR markers and Mendelian segregation of polymorphic bands was demonstrated. Among the 70 ISSR markers, simplex and duplex markers accounted for 70% and 15.7%, respectively. One linkage between two simplex ISSR markers was found. Thus, the ISSR markers could be suitable for finger-printing of cultivated sweetpotato and its related wild species and for constructing a linkage map of sweetpotato.