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Phoenix dactylifera L. is widely distributed and performs an important socioeconomic role in the south of Tunisia. The objective of this work was to evaluate different methods of estimating the diversity and genetic structure of Tunisian date palm subpopulations. Nine morphological traits and five SSR loci were used to study morphological and genetic diversity. Correlations between phenotypic and genetic distances were assessed. Analyses of variance of the morphological data reveal significant differences among subpopulations for all traits measured. The Mantel test shows that morphological variation is correlated with fruit maturity period (r = 0.161; p = 0.020). Analysis of molecular variance reveals significant genetic variation among fruit-consistency subpopulations (p < 0.05) and the Mantel test emphasises a correlation between genetic distance and fruit consistency (r = 0.110; p = 0.029). This information will be useful for germplasm collection, for conservation and for various date palm culture programmes in the Tunisian continental oases.