The phylogenetic relationships among 26 taxa of Brachycera (Diptera) beside one outgroup taxa of Nematocera were studied using cladistic analysis and one PCR-amplified DNA gene segments (28S). The cladistics analysis was applied by Mesquite program (using 216 multistate characters). The taxa were sampled from various areas of Egypt. The morphometric analysis and phenotypic features were determined for the all selected brachyceran taxa to reveal the phylogenetic relationships among them. The resulted cladogram showed high degree of variations and affinities (Similarity coeffeciant ranged from 53.8 to 97.8%).We have sequenced 205 bp of 28S gene of all taxa and the comparison of the nucleotide compositions found allows phylogenetic analyses of the suborder. The estimated net nucleotide sequence divergence among the taxa examined was found to range from 0.00 to 0.7% as an affinity. Cluster analysis based on both morphometric and sequence data showed less variation than morphometric data only (Similarity coeffeciant ranged from 60.6 to 82.5%) of the 27 taxa. The trees obtained from combined datasets (morphology/28S gene) using (Parsimony method) revealed the radiation of the basal lineages of the suborder.