Ziziphus spina-christi (Christ’s thorn jujube) is a culturally and economically significant indigenous fruit tree species in Sudan, particularly valued by rural communities and women for its nutritional and income-generating potential. Despite its widespread use and apparent diversity, systematic characterization of existing populations has been lacking. This study aimed to assess the morphological and genetic diversity of Z. spina-christi and explore indications of domestication in the Nuba Mountains. A total of 250 individual trees were sampled equally from agroforestry homegardens and adjacent forests across five locations. Morphometric traits of trees and fruits were recorded, and dried leaves were subjected to amplified fragment length polymorphism (AFLP) analysis. Statistical methods including Pearson’s correlation, t-tests, Mann-Whitney tests, and regression analyses were employed to identify significant relationships between traits and environmental factors. Results revealed substantial morphological diversity in traits such as stem diameter, fruit size, and fruit weight. Fruit size negatively correlated with both latitude and elevation. Trees in homegardens exhibited significantly larger stem diameters than those in forests, suggesting human selection. Regression analyses indicated that fruit size and dry weight were positively influenced by homegarden settings and canopy area, but negatively affected by elevation. Preliminary AFLP results revealed a distinct genetic differentiation between homegarden and forest populations, implying limited seed flow and different selection pressures. These findings suggest that Z. spina-christi in the Nuba Mountains is undergoing a process of semi-domestication driven by environmental conditions and human cultivation practices. This study provides a foundation for developing strategies for the local domestication and in situ conservation of this important species.