DNA Barcoding for Managing Blackberry Genetic Resources on Black Sea Coast (Russia)

Accurate species identification in blackberries (Rubus spp.) is difficult because of morphological similarity and frequent hybridization. We studied 56 wild accessions from the Sirius Federal Territory (Russia), representing coastal and foothill ecosystems of the Black Sea region. Multilocus DNA barcoding with the plastid rbcL gene and nuclear ITS1 and ITS2 regions revealed signals of hybridization and hidden diversity. The rbcL marker showed low variation, grouping most accessions into two clusters with several singletons, which limited its use for distinguishing species. In contrast, ITS1 and ITS2 showed higher variation, forming six clusters and eight singletons, and allowed for clear separation of taxa such as Rubus caesius L., R. irritans Focke, and R. amabilis Focke. Accession 3 carried a raspberry (closely to R. corchorifolius L.fil) plastid haplotype, pointing to a hybrid origin. We also found groups of nearby plants with identical mutations, which likely reflect clonal spread with fixed somatic changes or the persistence of recent hybrid lineages. At the same time, accessions collected up to 140 km apart did not form separate clusters, showing weak geographic structuring along the coast. The results demonstrate that multilocus barcoding can reveal not only species boundaries but also evolutionary processes among Rubus such as hybridization, clonal propagation, and early stages of speciation.