To face the increasing global climate change in water level and to achieve sustainable development of seed yield and oil yield production in the sesame crop, five cultivars of sesame, H 102/58 (P1), M2 A6 (P2), M1 A38 (P3), shandaweel3 (P4), and H 11/9 (P5), were employed to implement a half-diallel mating design. Studied cultivars and their hybrids were grown for evaluation under two water regimes: normal irrigation and drought stress, to study the growth, yield performance, heritability, heterosis, and combining ability. Results under normal and stress conditions showed that all the studied parents and their crosses had different distinct traits, especially P3, which had the highest mean value for most growth and yield traits. P3, P4, and P5 had the highest mean values for the 1000 seed weight trait, while P2 recorded the highest mean values for oil content percent. The superiority of crosses was detected by positive and significant values of specific combining ability (SCA) effects and heterosis in P1 ×P3, P2 ×P4, and P4 ×P5. Start codon targeted (SCoT) polymorphism is an effective method for fingerprinting and genetic diversity analysis. 126 amplified fragments with a 52% polymorphism of 15 Sesamum indicum L. genotypes were obtained in the current study utilizing 10 SCoT primers. In this instance, there were 66 polymorphic bands and 60 monomorphic bands. Genetic similarity ranged from 75% to 92%, according to the SCoT data analysis. The 15 genotypes of sesame examined were divided into two main clusters and two main sub-clusters using a dendrogram.