This study was conducted during 2015, 2016 and 2017 seasons at Antoniades Research Branch, Horticulture Research Institute, Ministry of Agriculture, Alexandria. The aim of the present study was to study the effects of different doses of gamma rays from Cobalt -60 on Cyperus alternifolius L. plants Rhizomes of Cyprus plants were irradiated with six doses of gamma irradiation (0.0,20, 40, 60, 80 and 100 Gray). The results showed that irradiation of rhizomes with 20 Gray caused the highest increase in sprouting percentage and  rate while the 100 gray treatment caused the lowest survival percentage in M1 –generation. The 100 Gray treatment caused the lowest significant decrease in all studied vegetative growth parameters in M1 and M2 generations.  As for the vegetative growth abnormalities, the application of gamma rays at the dose of 100 Gray resulted in dwarf plants in M1 and M2 generations. While using 40 Gray caused 3 regenerated small plants from leaves internode and  60  Gray caused 1 regenerated small plant from leaves internodes in M1 –generation , this abnormality disappeared in the M2 –generation. Application of  restriction fragment length polymorphism (RFLP) analysis  on three Cyperus alternifolius generations stressed  with  60 and 100 Gray  comparing with un-irradiated Cyperus alternifolius cleared that radiation dose (100 Gray) reflected the dramatic increase of genetic polymorphism and first generation reflected the highest radiation influence on polymorphism.