A Population of four canola variants and their derived gamma rays- induced mutant lines were used to examine genetic diversity among them based on ISSR- PCR analysis. The study was extended to find if any ISSR variants were related to seed yield and oil percentage. The evaluation of mutant lines for three consecutive years at two different locations (Inshas and Ras Suder) for seed yield and its attributes showed that lines 92 and 63 possessed highly significant performance than the other mutants and their parents for seed yield /plant in both locations. Thirteen ISSR primers were used in this study. All primers generated 1064 DNA bands, with an average of 81.84 bands per primer, 980 bands of them were polymorphic. All primers detected unique bands (84 bands); no monomorphic bands were observed in this study. The profiles generated by primer ISSR2 contained the highest percentage of polymorphic bands (97.16%). Results showed that lines 92 and 63 generated from Evita and Pactol cultivars possessed the highest seed yield /plant in Inshas and Ras Sudr locations, respectively. Those lines also showed the highest number of unique, bands which could be used as candidate molecular markers for high seed yield. Data showed that line 92 and 63 are excellent performing new genotypes which could be introduced in breeding program to obtain new Egyptian canola varieties. Theprimers ISSR2, ISSR4, ISSR6 and ISSR9 could be considered specific primer for canola genotypes in searching for the discovery of molecular markers associated high seed yield/ plant and consequently oil percentage.