Aim and Background. Gladiolus italicus species are exposed to genetic erosion and even destruction. On the other hand, these species can be potentially useful for breeding and agriculture. Moreover, the diversity of germplasm is not sufficient. Therefore, the collection, preservation and study of genetic diversity on Gladiolus species are important and essential.

Materials and Methods. According to Flora of Iran books, different regions of the country were visited. Three populations of Gladiolus species from different area namely Alamout, Mahalat and Mashhad were collected and their karyotype characteristics were recorded .Three plants of each population was randomly selected for further studying. The root tips pretreated with 8-hydroxyquinolin for 3/5 hours, then fixed in Former solution 1:3 ethanol and acetic acid for 24 hours , followed by being stained with hematoxylin. Images of ten appropriate metaphase plates with clearly chromosomes morphology were then captured.

Conclusion. Karyotype symmetry, number of SAT and chromosomes differed significantly. chromomatin length as well as length of chromosome arms were also differed. Therefore, medium-chromosome was designated as diploid.