The use of barcoding sequences for phylogenetic assessment in Meliaceae from fragments of rainforest in Indonesia

DNA barcoding is a growing concept in modern biological science which is a technique of sequencing extracted genomes for exploration and taxonomic clarification of species with high confidentiality in a short period.
This research aims to evaluate the feasibility of DNA barcodes for the identification of Meliaceae species, to compare morphological identification with DNA barcoding assignments, and to reconstruct the phylogenetic relationship of Meliaceae family using matK, rbcL, and ITS2 barcodes. The specimens were collected from 32 (50*50m) core plots in the Jambi province, Sumatra, Indonesia which was dried in silica-gel and stored in different herbariums of Indonesia. The herbarium vouchers were thoroughly compared to the reliable reference vouchers from Indonesia and a total of 87 Meliaceae species were morphologically identified by the taxonomists. All the laboratory experiments were carried out in the Department of Forest Genetics and Forest Tree Breeding, Göttingen, Germany. The sequences were edited and aligned using Codoncode AlignerTM software. The rbcL (95.40 %) fragment has the higher PCR sequencing success rate compared to the matK (91.95%) and ITS2 (63.21%) region. The homologous sequences for each samples were retrieved from NCBI GenBank. The nBLAST supported ITS2 marker as an effective marker to find higher resemblance of molecular and morphological identification of sample to the species level (23.26 %) and genus level (74.42 %) whereas matK and rbcL were only efficient for lower discrimination level i.e. genus and family level respectively. The genetic distance was estimated using MEGA7 software. The highest overall mean genetic distance was estimated by nuclear ITS2 (0.082) region and the significant barcode gaps were observed in ITS2+matK and ITS2+rbcL region. The phylogenetic trees constructed using Maximum Likelihood and the Bayesian inference methods indicated Meliaceae has a sister relationship to family Rutaceae and subfamily Melioideae was recovered a monophyletic clade. Most tribes in Melioideae appeared as monophyletic lineages i.e. Aglaieae Sandoriceae, Meliaeae, and Turraeeae whereas Guareeae and Trichileae were recovered as a non-monophyletic clades. The phylogenetic assessment carried out using the concatenated sequences based on the combined matrix of ITS2+matK, ITS2+rbcL and ITS2+matK+rbcL provide successful phylogenetic analyses at the species level with strong clade support in Meliaceae family and these markers are also effective for a barcode gap analysis. So, individual ITS2 along with the combination with plastid regions serves as an excellent barcode in the Meliaceae family from Sumatra, Indonesia.