Abstract
DNA barcoding is used as a universal tool for delimiting species boundaries in taxonomically challenging groups, with different plastid and nuclear regions (rbcL, matK, ITS and psbA-trnH) being recommended as primary DNA barcodes for plants. We evaluated the feasibility of using these regions in the species-rich genus Terminalia, which exhibits various overlapping morphotypes with pantropical distribution, owing to its complex taxonomy. Terminalia bellerica and T. chebula are ingredients of the famous Ayurvedic Rasayana formulation Triphala, used for detoxification and rejuvenation. High demand for extracted phytochemicals as well as the high trade value of several species renders mandatory the need for the correct identification of traded plant material. Three different analytical methods with single and multilocus barcoding regions were tested to develop a DNA barcode reference library from 222 individuals representing 41 Terminalia species. All the single barcodes tested had a lower discriminatory power than the multilocus regions, and the combination of matK+ITS had the highest resolution rate (94.44%). The average intra-specific variations (0.0188±0.0019) were less than the distance to the nearest neighbour (0.106±0.009) with matK and ITS. Distance-based Neighbour Joining analysis outperformed the character-based Maximum Parsimony method in the identification of traded species such as T. arjuna, T. chebula and T. tomentosa, which are prone to adulteration. rbcL was shown to be a highly conservative region with only 3.45% variability between all of the sequences. The recommended barcode combination, rbcL+matK, failed to perform in the genus Terminalia. Considering the complexity of resolution observed with single regions, the present study proposes the combination of matK+ITS as the most successful barcode in Terminalia.
Highlights
IntroductionThe genus Terminalia Linn. (the second largest genus in the Combretaceae) comprises approximately 200 species of trees and shrubs [1, 2] with pantropical distribution [3, 4]
The genus Terminalia Linn. comprises approximately 200 species of trees and shrubs [1, 2] with pantropical distribution [3, 4]
The primers for internal transcribed spacer (ITS) used in the study lie in the conserved flanking regions of 18S and 26S, so the sequences were trimmed to boundaries of the ITS1, 5.8S and ITS2 region. rbcL was found to be highly conserved (95.9% identical sites) with the lowest number of variable sites (22/636), while matK and ITS showed the highest variability and length variation
Summary
The genus Terminalia Linn. (the second largest genus in the Combretaceae) comprises approximately 200 species of trees and shrubs [1, 2] with pantropical distribution [3, 4]. (the second largest genus in the Combretaceae) comprises approximately 200 species of trees and shrubs [1, 2] with pantropical distribution [3, 4]. In India, nearly 24 species have been reported from various regions [5]. DNA barcode reference library for genus Terminalia. All other data supporting the results of this article are available online in BOLD database under the project CRCBT – "DNA Barcoding in genus Terminalia". The sequence alignments generated and analysed during the current study are available as Supporting files
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