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The sunflower genus, Helianthus L. includes around 50 species native to the North America and appeared in Iran about one century ago. The purpose of this study is classification of all populations of H. tuberosus L. in 3 Northen Provinces (Guilan, Mazandaran and Golestan) of Iran by palynological investigations. We used Scanning Electron Microscopy (SEM) for 6 Populations. We observed some interesting differences between pollen grains belonging to eastern and western populations of the area. Pollen grains of H. tuberosus are Echinate. The populations of Guilan and Mazandaran provinces are 3-Colporate, but the populations of Golestan are Porate, without any Colpi and Opercul. In studied area, has been observed that the pollens from east to west, the Pores were changed to Colpies. The smallest pollen grains were observed in Tirtash and largest in Somehsara ranging from 34.77 to 24.71 μm in size. Length of the echinus and their distance from each other was equal in all populations. Statistical analysis including Principal Component Analysis and cluster analaysis showed significant differentiation between studied populations. According to the diagram and the table of variances of pollen traits, it seems that two main components have the main role in total variance. The first and the second main components explain 78.71 and 10.37 percent of total variances respectively.