In the present study, the phenological and genetic differences/similarities of 95 walnut genotypes were considered. These genotypes had been propagated sexually from the open pollination of the maternal cultivar ‘Maraş 12’. This study examined the timing of several phenological features like bud break, leafing, yellowing of the leaves and defoliation in the mother plant and in its progenies. A total of 6 ISSR primers were used for determining genetic differences among the genotypes. The differences between the maternal cultivar and the offspring genotypes included a difference of 97.89% in the bud-break date, 96.84% in the leafing date, 100% in the yellowing date of the leaves and 94.73% in the defoliation date. On average, the phenological variation stood at 97.36%. PCA analysis also confirmed the presence of genetic diversity-based phenological markers in our walnut seedling collection with PC1 58.27%, PC2 27.16%, PC3 12.16%, and PC4 2.40% of the total variation. A total of 38 bands were obtained in molecular analyses. These bands were largely polymorphic (89.47%). While the average number of alleles was 5.66, Polymorphism Information Content (PIC) varied between genotypes from 0.47 to 0.95, and the average PIC value was 0.64. The UPGMA dendrogram divided the genotypes into two main clusters and the similarity coefficient of the genotypes varied between 0.68 and 0.96. In sum, it was found that open-pollinated walnut genotypes had high degrees of phenotypic and genetic variation in comparison with their maternal cultivar ‘Maraş 12’.