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Melilotus is an important genus of legume plants and an herbage with excellent nitrogen fixation; it can tolerate extreme environmental conditions and possesses important medicinal value. However, there is limited genetic information about the genus; thus, we analysed four chloroplast loci (rbcL, matK, psbA-trnH and trnL-F) and one nuclear region (ITS) to determine the genetic diversity of 199 accessions from 18 Melilotus species. The rbcL and matK sequences were highly conserved, whereas the trnL-F and ITS sequences contained variable and parsimony-informative sites. In our analyses of the single and combined regions, we calculated the pairwise distance, haplotype and nucleotide diversity and gaps and then constructed phylogenetic trees to assess the genetic diversity, and our results revealed significant variations among the different accessions. The genetic distance values were between zero and nine, and based on the combined regions, the highest frequency value was approximately four. Melilotus showed high haplotype and nucleotide diversity, particularly in the ITS sequences, with values of 0.86 and 0.0087, respectively. The single ITS sequence, psbA-trnH, and the combined matK+rbcL+trnL-F (MRT) and matK+rbcL+psbA-trnH+trnL-F+ITS (MRPTI) regions showed interspecific variation in the gap analysis. Phylogenetic trees calculated using ITS, psbA-trnH and MRPTI sequences indicated distinct genetic relationship in 18 species, and these species could be divided into two groups. By determining the genetic diversity of plants, we can evaluate the genetic relationships among species and accessions, providing a basis for preserving and utilizing the genetic resources of Melilotus. [ABSTRACT FROM AUTHOR]

Melilotus, an annual or biennial herb, belongs to the tribe Trifolieae (Leguminosae) and consists of 19 species. As an important green manure crop, diverse Melilotus species have different values as feed and medicine. To identify different Melilotus species, we examined the efficiency of five candidate regions as barcodes, including the internal transcribed spacer (ITS) and two chloroplast loci, rbcL and matK, and two non-coding loci, trnH-psbA and trnL-F. In total, 198 individuals from 98 accessions representing 18 Melilotus species were sequenced for these five potential barcodes. Based on inter-specific divergence, we analysed sequences and confirmed that each candidate barcode was able to identify some of the 18 species. The resolution of a single barcode and its combinations ranged from 33.33% to 88.89%. Analysis of pairwise distances showed that matK+rbcL+trnL-F+trnH-psbA+ITS (MRTPI) had the greatest value and rbcL the least. Barcode gap values and similarity value analyses confirmed these trends. The results indicated that an ITS region, successfully identifying 13 of 18 species, was the most appropriate single barcode and that the combination of all five potential barcodes identified 16 of the 18 species. We conclude that MRTPI is the most effective tool for Melilotus species identification. Taking full advantage of the barcode system, a clear taxonomic relationship can be applied to identify Melilotus species and enhance their practical production. [ABSTRACT FROM AUTHOR]

Melilotus comprises 19 species, while the phylogenetic relationships between species remain unclear. In the present work, three chloroplast genes, rbcL, matK, trnL-F, and one nuclear region, ITS (internal transcribed spacer) belonging to 48 populations of 18 species of Melilotus were sequenced and phylogenetic trees were constructed to study their interspecific relationships. Based on the phylogenetic tree generated in this study using rbcL analysis, the Melilotus genus is clearly monophyletic in the legume family. Both Bayesian and maximum-parsimony approaches were used to analyze the data. The nrDNA ITS provided more informative characteristics (9.8%) than cpDNA (3.0%). Melilotus contains two closely related groups, clade I and clade II. M. spicatus, M. indicus and M. segetalis have a close relationship. M. infestus, M. siculus and M. sulcatus are closely related. The comparing between molecular phylogeny and flower color classification in Melilotus showed that the flower color is not much informative for phylogenetics of this genus. [ABSTRACT FROM AUTHOR]