This study conducted to determine relationship among and within Iranian Helichrysum species (Asteraceae). In this study based on ISSR markers, the highest percentage of ISSR loci polymorphism (54.7%) occurred in H. armenium. The highest gene diversity over loci (1.224), Shannon’s Information Index (0.224%) and Expected Heterozygosity (0.142%) occurred in H. armenium (0.18) and the lowest of these parameters (0%) were observed in H. araxinum, H. graveolens, H. persicum and H. psychrophilum. The highest genetic similarity occurred between H. armenium and H. rubicundum (0.989), while the lowest was between H. polyphyllum and H. graveolens (0.213). The analysis of molecular variance (AMOVA), showed significant genetic variation among (24%) and within (76%) species. In morphological analysis traits such as indumentum, resting bud, achene length, achenial papillae, dimension of receptacle and form and apex of phyllaries were main diagnostic features. Results obtained from the morphological cluster were greatly consistent with the molecular data, to elucidating taxonomic relationships, as well as both attributed the higher diversity in H. armenium and H. rubicundum in comparison with other species and also indicated that H. persicum is a member of H. oocephalum species. Totally we confirmed the presence of 18 species in Iran.