Evaluation of chloroplast DNA markers for distinguishing Phalaenopsis species

• We evaluated 27 cpDNA regions to identify markers for distinguishing moth orchids.
• More than 62% of cpDNA markers were highly variable at the species level.
• The rps16-trnQ region showed the best discrimatory power.
• The maternal inheritance mode of cpDNA in moth orchids was predominated.
• The non-monophyletic relationship of cpDNA was revealed below Phalaenopsis genus.

Comparative analysis of chloroplast DNA (cpDNA) between two endemic moth orchids in Taiwan, Phalaenopsis aphrodite subsp. formosana and P. equestris, previously revealed many evolutionary hotspots. In this study, we explored 27 pre-screened cpDNA regions to evaluate molecular markers for distinguishing Phalaenopsis species. Fifteen cpDNA markers were highly variable among moth orchids, with polymorphic information contents ≥8.0. The rps16-trnQ marker showed the best discrimatory power which 15 endemic moth orchids could be successfully separated into 13 groups. From the sequences of the seven selected cpDNA regions, the rps16-trnQ intergenic spacer was the best DNA barcode which 19 endemic moth orchids could be completely distinguished. The petN-psbM intergenic spacer could be better used as a DNA barcode to differentiate P. aphrodite subsp. formosana and P. amabilis. Additionally, the inheritance mode of cpDNA in moth orchids was maternally predominated. Phylogenetic analysis revealed non-monophyletic relationships below the Phalaenopsis genus level. In summary, we have revealed a set of cpDNA markers that could be used for identification and phylogenetic study of Phalaenopsis orchids.