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Abstract Background Electrophoretic SDS-PAGE and SCoT-PCR techniques were used to study the fingerprints and genetic diversity of six Egyptian soybean (Glycine max L.) genotypes (Giza111, Giza21, Giza82, Giza35, Giza22, and Giza83). Results SDS-PAGE revealed a total of 23 bands with a polymorphism 30.43%. The maximum number of bands (22) was recorded in Giza111 and Giza82, while the minimum number of bands (18) was recorded in Giza35. The highest similarity index for SDS- PAGE (0.909) was observed between Giza111 and Giza82, while the lowest similarity index (0.714) was recorded between Giza35 and Giza83. SCoT technique was performed by using 11 primers to generate reproducible polymorphic products. The total number of bands was 106, from them 52 polymorphic and 54 monomorphic bands with a polymorphism 49.11%. The dendrogram of both SDS-PAGE and SCoT markers revealed two main clusters; the first cluster includes the cultivars Giza111and Giza21, while the second cluster includes the cultivars Giza82, Giza35, Giza22, and Giza83. The second group was further subdivided into two subgroups; the first subgroup includes cultivars (Giza82 and Giza35) while the second subgroup includes cultivars Giza22 and Giza83. Conclusion The markers used in this study created a number of polymorphic bands among the different cultivars that can be utilized as molecular markers for their differentiation. The obtained data indicated that SCoT and SDS-PAGE techniques could be used efficiently for identification and differentiation of the selected genotypes.