The gland struc- ture was examined by direct field observations. Taxonomically important characters of glands were observed and measured: size, texture, shape, color, and horns. For species out of Iran herbarium materi- als were studied. Seed characteristics were examined using scanning electron microscopy (SEM) as well as dissecting light microscopy. Significant features are: seed size, seed shape, presence of caruncle, shape of caruncle, and seed surface ornamentation. A phylogenetic study using Maximum Parsimony (MP) and Bayesian Inference (BI) was performed based on sequences of nuclear DNA internal transcribed spacers (ITS) for selected species representing the main clades known in Euphorbia and with special focus on the species distributed in Iranian highlands. ITS sequences for 20 accessions representing 19 species are provided for the first time, and 48 accessions of 47 species were used from GenBank. The topologies of both analyses were congruent. The results indicate: (1) four main clades with high supports in sub- gen. Esula which are appropriate to be recognized at sectional rank. (2) E. larica is nested within clade A including few members of subgen. Rhizanthium and is closely related to sect. Balsamis, which is suggested here to be transferred from subgen. Esula into subgen. Rhizanthium. (3) E. osyridea of the monotypic sub- sect. Osyrideae is closely related to E. buhsei and to the members of sect. Esula. Tracing morphological characters on the phylogenetic tree shows that several morphological characters, such as seed ornamen- tation applied in previous subgeneric classification of the subgen. Esula, are homoplasious, but the gland structure and capsule surface characters are more reliable for classification purposes.