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The steno-endemic species of genus  Decalepis  are highly threatened by destructive wild harvesting. The medicinally important fleshy tuberous roots of  Decalepis hamiltonii  are traded as substitute, to meet the international market demand of  Hemidesmus indicus . In addition, the tuberous roots of all three species of  Decalepis  possess similar exudates and texture, which challenges the ability of conventional techniques alone to perform accurate species authentication. This study was undertaken to generate DNA barcodes that could be utilized in monitoring and curtailing the illegal trade of these endangered species. The DNA barcode reference library was developed in BOLD database platform for candidate barcodes  rbcL ,  matK ,  psbA-trnH ,  ITS  and  ITS2 . The average intra-specific variations (0–0.27%) were less than the distance to nearest neighbour (0.4–11.67%) with  matK  and  ITS . Anchoring the coding region  rbcL  in multigene tiered approach, the combination  rbcL  +  matK  +  ITS  yielded 100% species resolution, using the least number of loci combinations either with PAUP or BLOG methods to support a character-based approach. Species-specific SNP position (230 bp) in the  matK  region that is characteristic of  D .  hamiltonii  could be used to design specific assays, enhancing its applicability for direct use in CITES enforcement for distinguishing it from  H .  indicus .

Character-based DNA barcoding for authentication and conservation of IUCN Red listed threatened species of genus Decalepis (Apocynaceae)