Internal Transcribed Spacer (ITS) Fails Barcoding of the Genus Neotinea Rchb.f. (Orchidaceae)

Internal Transcribed Spacer (ITS) is one of the most used barcoding regions for molecular phylogenetics and orchids. Our aim in this study to test reliability ITS on closely related Neotinea spp., including tridentata, ustulata subsp. aestivalis, by comparing it accD-psaI intergenic spacer plastid DNA. Both were amplified specific primer sets sequenced. Phylogenetic trees regenerated using Maximum Parsimony approach. The results showed that separated some N. tridentata samples Turkish, Greek, Hungarian Croatian from others phylogenetic due incomplete lineage sorting. In contrast ITS, marker could successfully separate according a priori species classification. findings refer hybridisation story between ustulata. We propose not use sequences directly as barcode reconstruct phylogeny group. Instead, inclusion other nuclear such LFY, ADH, etc., or utilisation whole genome sequencing give better results.