The genus Linum L. is an important plant genus as it contains the species with economic values and particularly Linum usitatissimum L. that is source of fibre and linseed oil. This genus contains 230 species throughout the world and has about 22 species in Iran. Little is known about Linum species relationship and phylogeny. Therefore, the aim of present study was molecular phylogenetic investigation of the Linum species growing in Iran and to present data on their biogeography. We used both ITS and chloroplast DNA sequences (psbA-trnHGUG region) for inferring the species phylogeny and relationship. We also used cpDNA for inferring the species time of divergence and with ISSR markers to identify the path of species distribution in the country. The phylogenetic trees obtained for both ITS and cpDNA sequences were almost congruent. NeighborNet diagram and BEAST tree based on Bayesian method separated the outgroup species Hugonia and Anisadenia from the other species studied. The subspecies studied in Linum macronicum were placed close to each other and along with L. corymbulosum comprised a separate clade. The clades obtained showed divergence time between 5–20 mya. The present study revealed that the species of the sect. Linum are monophyletic, while members of the sections Linastrum and Syllinum are intermixed and seem to be paraphyletic.