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Due to the economic importance of the date palm in Tunisia, we attempt to develop microsatellite markers for the genetic characterisation and identification of cultivars. This characterisation will enable us to obtain an extensive understanding of the local genetic diversity and its distribution. Fourteen microsatellite loci of Phoenix dactylifera L. were tested to examine the genetic diversity in the Tunisian date palm germplasm: 74 female and 27 male cultivars were represented. A total of 136 alleles and 311 genotypes were detected based on 14 loci microsatellite. A high level of polymorphism was detected at the DNA level. The 101 date palms from three main oases exhibited a high degree of generic diversity and were structured independently of their geographical origin and sex. All the local date palm accessions were successfully fingerprinted and easily distinguished based on only five loci. The possibility of using microsatellites in the large-scale molecular labelling of offshoots and in vitro plantlets and in the certification of plant material is discussed. (c) 2012 Elsevier B.V. All rights reserved.