Iran with a diverse climate and rich herbal diversity is one of the richest countries in natural genetic resources. Iran is also the main origin and one of the most important centers of Astragal species diversity. Astragalus gossypinus Fischer which produce gum tragacanth, belongs to papilionacae and mainly spread in Asia. This species distributed in Iran, Iraq, Lebanon and syria. The most important habitats of gum producer Astragalus species are at montain ranges of IranoTooran region (especially Alborz and Zagros heights). Isfahan province have maximum number of Astragal species in Iran (154 species). Astragalus habitats in Isfahan province cover 31.1 percent of its area. Astragalus gossypinus mostly studied ecologically and morphologically. In this study, genetic attributes of Astragalus gossypinus populations in western rangelands of Isfahan province has been investigated and compared. The results of such studies could use in interbreeding, production and increase of Astragalus individuals with high quality gum tragacanth, reclamation of destructed habitats or amplification of Astragalus habitats. To do so, Astragalus gossypinus Fischer samples collected from six rangeland sites from its habitats in Isfahan province, including: Rahmat abad (Khansar), Ghale jamal (Golpayegan), Betlijeh Hendokosh (Fereidan), mousa abad (Tiran Karvan) and Tang bidakan (Lenjan). Genomic DNA was extracted using CTAB method and quality of extracted DNA was checked on electrophorus gel. DNA density measured by spektrofotometry method. After assuring the quality of DNA, DNA fragment of CNGC4 gene was amplified and the PCR product was checked on agaroze gel. The resulted product was sequenced and aligned using CLC Main Workbench5 program. For amplification, two primers designed by Oligo6 software. The PCR reaction has been done on a part of CNGC4 gene. The results of gene allignment of CNGC4 showed a genetic similarity between Astragalus gossypinus populations. Regarding genetic similarity, Astragalus gossypinus populations considered as one genetype. The genom of Astragalus gossypinus also compared with other Astragal species. The results of cluster analysis and UPGMA method indicated the existence of three major groups between Astragal species. According to these results and previous studies about quantity and quality of gum tragacanth products of this species, it can be concluded that since Astragalus gossypinus has a narrow ecological niche and resistance range, the difference in quantitative and qualitative attributes of its gum tragacanth in different habitats are caused from environmental conditions affecting physiological and functional life of Astragalus gossypinus and may not be attributed to genetic diversity of this species. Keywords: Astragalus gossypinus , Genetic diversity, DNA, PCR, Allignment, Cluster analysis, habitat, Isfahan province.