In this study, 15 simple sequence repeat (SSR) markers were used for genetic diversity analysis of 45 almond accessions, which included 25 local cultivars from La Palma Island and three other commercial cultivars. A total of 110 amplification fragments were produced, with an average value of 7.9 alleles per locus. Twelve of the SSR markers can be considered as highly informative, with values of expected heterozygosity and power of discrimination above 0.5 and 0.8, respectively. Due to cases of synonymy and homonymy, 37 different genetic profiles were obtained, with the homonymy of the soft-shell varieties known as ‘Mollar’ being the most significant. Cluster analysis identified four groups within the accessions. One of these groups exclusively consisted of the two commercial cultivars ‘Guara’ and ‘Ferraduel’. The other commercial cultivar used in the study, ‘Desmayo Largueta’, was in a cluster with three cultivars from the same locality. The analysis of molecular variance revealed that the within-localities component accounts for most of the total variation, suggesting that La Palma almond cultivars did not originate independently in different parts of the island. The results of the study reveal the genetic singularity of La Palma almond cultivars and the genetic diversity among them.