Molecular markers ISSR were used to analyse γ-irradiated mutants of amaranth Ficha cultivar and K-433 hybrid; tested six microsatellite -primed PCR markers. First analysis of gamma-irradiated amaranth mutant lines was done using the ISSR. In the study, primers that distinguish the mutant lines from control plant were reported as well as primer with the differentiation ability for all analysed mutant lines. The very specific changes in the mutant lines´ non-coding regions based on inter microsatellite length polymorphism were analysed. Mutant lines of the Ficha cultivar (C15, C26, C27, C82, C236) shared a genetic dissimilarity of 0.16 and their ISSR profiles were more similar to the Ficha than those of K-433 hybrid mutant lines. The K-433 mutant lines (D54, D279, D282) shared genetic dissimilarity of 0.22 but are more distinct to their control plant as a whole, as those of the Ficha mutant lines. Different ISSR fingerprints patters of the mutant lines when compared to the Ficha cultivar and K-433 hybrid ISSR specific profiles may be part of a consequence of the complex response of the intergenic space of mutant lines to the gamma-radiance.