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How far advanced is the DNA-based identification of the BELFRIT-List?
Abstract
The sector of botanicals is characterized by the huge number of species found in products on the market. Some governments issue positive and negative lists of plants in order to create legal certainty and to increase safety of those products. The most advanced positive list in Europe is the BELFRIT-List with 991 species in 594 genera, co-ordinate between Belgium, France and Italy. DNA-based methods for the identification of botanicals supplement nowadays the other identification methods based on morphology or phytochemistry. Molecular phylogenetic and population studies are helpful tools to develop a specific DNA-based method. In this contribution, the BELFRIT-List was reviewed for the availability of either a DNA-based identification method or molecular phylogenetic information. For 286 genera (48% of the genera), no helpful information could be found. Two hundred and thirty eight references with the intention to identify species by DNA demonstrate the already advanced field of this method. Such new methods are not developed systematically for all species of such lists, but more on a case-by-case approach for species difficult to identify. Therefore, the high number of papers already dealing with this method or bringing valuable information for their development should encourage combined efforts in standardizing validation.
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Hybridization plays a major role in speciation. However, hybridization and reticulate evolution in general are poorly understood in tree species because genetic documentation is often missing. Analyses of biparentally inherited gene regions allow detection of reticulate signals. Multicopy and single‐copy nuclear markers may show significant intraindividual variability owing to reticulation processes. Naturally, such processes induce incompatible phylogenetic signal resulting in incongruent genealogies. Data from three nuclear markers, two multicopy nrDNA spacers, and the single‐copy 2nd intron of the LEAFY gene, mirror ancient and recent horizontal gene flow in plane trees (Platanus). In addition to previously assembled data from the internal transcribed spacers (ITS) of the 35S ribosomal DNA, we found atypical 5S rDNA intergenic spacer sequences (5S‐IGS) causing significant intra‐ and interindividual polymorphism, and a conspicuous LEAFY intron dimorphism. A detailed framework of reticulate molecular evolution of Platanus can be erected using splits graphs based on distances between cloned sequences or individuals, and competing topologies. Two hundred and sixty‐one 5S‐IGS sequences and LEAFY genotyping of 71 individuals via sequence analysis and PCR‐RFLP support a previous ITS study (including pseudogenous and non‐pseudogenous variants) suggesting that the modern North American taxa P. rzedowskii and P. occidentalis var. palmeri are the result of ancient hybridization. Platanus occidentalis var. palmeri requires taxonomic revision and is provisionally treated at species rank.
This project examines the relationships within the genus Zea using complete plastid genomes (plastomes). While Zea mays has been well studied, congeneric species have yet to be as thoroughly examined. For this study four complete plastomes and a fifth nearly complete plastome were sequenced in the five species (Zea diploperennis, Zea perennis, Zea luxurians, Zea nicaraguensis, and Zea mays subsp. huehuetenangensis) by Sanger or next-generation methods. An analysis of the microstructural changes, such as inversions, insertion or deletion mutations (indels) and determination of their frequencies were performed for the complete plastomes. It was determined that 193 indels and 15 inversions occurred across the examined plastomes of Zea. Tandem repeat indels were the most common type of microstructural change observed. Divergence times were estimated using a noncorrelated relaxed clock method. Divergence dates for specific nodes relative to Zea were calculated to fall between 38,000 years before present (YBP) for the subspecies included in this study and 23,000 YBP for section Luxuriantes included in this study. The stem lineage of all Zea species was calculated to have diverged at 176,000 YBP. The calculated mutation rates for the genus fell within the range of 1.7E−8 to 3.5E−8 microstructural changes per site per year. These rates of change are not uniform, despite the close relationships of taxa in this study. Phylogenomic analyses using full plastome alignments were also conducted to compare tree topologies from different types of mutations. In most cases, the previous work examining Zea mitochondrial and nuclear data was confirmed.
Several taxonomic and phylogenetic studies have hypothesized polyphyly within Pueraria DC., a genus comprising 19 species (24 with varieties) including the highly invasive Pueraria montana var. lobata (Kudzu) introduced to the U.S.A. about 150 years ago. Previous efforts to investigate monophyly of the genus have been hampered by limited taxon sampling or a lack of comprehensive evolutionary context that would enable definitive taxonomic associations. This work presents a comprehensive phylogenetic investigation of Pueraria within the context of tribe Phaseoleae (Leguminosae). Polyphyly was found to be more extensive than previously thought, with five distinct lineages spread across the tribe and spanning over 25 mya of divergence strongly supported by two chloroplast and one nuclear marker, AS2, presented here as a phylogenetic marker for the first time. Our phylogenies support taxonomic revisions to rectify polyphyly within Pueraria, including the resurrection of Neustanthus, moving one species to Teyleria, and the creation of two new genera, Haymondia and Toxicopueraria (taxonomic revisions published elsewhere).
Recently, commercial interest in Physalis species has grown worldwide due to their high nutritional value, edible fruit, and potential medicinal properties. However, many Physalis species have similar shapes and are easily confused, and consequently the phylogenetic relationships between Physalis species are poorly understood. This hinders their safe utilization and genetic resource conservation. In this study, the nuclear ribosomal ITS2 region was used to identify species and phylogenetically examine Physalis. Eighty-six ITS2 regions from 45 Physalis species were analyzed. The ITS2 sequences were aligned using Clustal W and genetic distances were calculated using MEGA V6.0. The results showed that ITS2 regions have significant intra- and inter-specific divergences, obvious barcoding gaps, and higher species discrimination rates (82.2% for both the BLASTA1 and nearest distance methods). In addition, the secondary structure of ITS2 provided another way to differentiate species. Cluster analysis based on ITS2 regions largely concurred with the relationships among Physalis species established by many previous molecular analyses, and showed that most sections of Physalis appear to be polyphyletic. Our results demonstrated that ITS2 can be used as an efficient and powerful marker in the identification and phylogenetic study of Physalis species. The technique provides a scientific basis for the conservation of Physalis plants and for utilization of resources.
In the past decades, the use of traditional medicine has increased globally, leading to a booming herbal medicine and dietary supplement industry. The increased popularity of herbal products has led to a rise in demand for botanical raw materials. Accurate identification of medicinal herbs is a legal requirement in most countries and prerequisite for delivering a quality product that meets consumer expectations. Traditional identification methods include botanical taxonomy, macroscopic and microscopic examination, and chemical methods. Advances in the identification of biological species using DNA-based techniques have led to the development of a DNA marker-based platform for authentication of plant materials. DNA barcoding, in particular, has been proposed as a means to identify herbal ingredients and to detect adulteration. However, general barcoding techniques using universal primers have been shown to provide mixed results with regard to data accuracy. Further technological advances such as mini-barcodes, digital polymerase chain reaction, and next generation sequencing provide additional tools for the authentication of herbs, and may be successful in identifying processed ingredients used in finished herbal products. This review gives an overview on the strengths and limitations of DNA barcoding techniques for botanical ingredient identification. Based on the available information, we do not recommend the use of universal primers for DNA barcoding of processed plant material as a sole means of species identification, but suggest an approach combining DNA-based methods using genus- or species-specific primers, chemical analysis, and microscopic and macroscopic methods for the successful authentication of botanical ingredients used in the herbal dietary supplement industry.
Georg Thieme Verlag KG Stuttgart · New York.
Quercus is considered economically and ecologically one of the most important genera in the Northern Hemisphere. Oaks are taxonomically perplexing because of shared interspecific morphological traits and intraspecific morphological variation, which are mainly attributed to hybridization. Universal plastid markers cannot provide a sufficient number of variable sites to explore the phylogeny of this genus, and chloroplast genome-scale data have proven to be useful in resolving intractable phylogenetic relationships. In this study, the complete chloroplast genomes of four Quercus species were sequenced, and one published chloroplast genome of Quercus baronii was retrieved for comparative analyses. The five chloroplast genomes ranged from 161,072 bp (Q. baronii) to 161,237 bp (Q. dolicholepis) in length, and their gene organization and order, and GC content, were similar to those of other Fagaceae species. We analyzed nucleotide substitutions, indels, and repeats in the chloroplast genomes, and found 19 relatively highly variable regions that will potentially provide plastid markers for further taxonomic and phylogenetic studies within Quercus. We observed that four genes (ndhA, ndhK, petA, and ycf1) were subject to positive selection. The phylogenetic relationships of the Quercus species inferred from the chloroplast genomes obtained moderate-to-high support, indicating that chloroplast genome data may be useful in resolving relationships in this genus.
To detect and avoid illegal logging of valuable tree species, identification methods for the origin of timber are necessary. We used next-generation sequencing to identify chloroplast genome regions that differentiate the origin of white oaks from the three continents; Asia, Europe, and North America. By using the chloroplast genome of Asian Q. mongolica as a reference, we identified 861 variant sites (672 single nucleotide polymorphisms (SNPs); 189 insertion/deletion (indel) polymorphism) from representative species of three continents (Q. mongolica from Asia; Q. petraea and Q. robur from Europe; Q. alba from North America), and we identified additional chloroplast polymorphisms in pools of 20 individuals each from Q. mongolica (789 variant sites) and Q. robur (346 variant sites). Genome sequences were screened for indels to develop markers that identify continental origin of oak species, and that can be easily evaluated using a variety of detection methods. We identified five indels and one SNP that reliably identify continent-of-origin, based on evaluations of up to 1078 individuals representing 13 white oak species and three continents. Due to the size of length polymorphisms revealed, this marker set can be visualized using capillary electrophoresis or high resolution gel (acrylamide or agarose) electrophoresis. With these markers, we provide the wood trading market with an instrument to comply with the U.S. and European laws that require timber companies to avoid the trade of illegally harvested timber.
Scandiceae subtribe Daucinae encompasses umbellifers that have fruits with prominent secondary ridges projecting into wings (former tribe Laserpitieae) or spines (former tribe Caucalideae pro parte). It comprises several economically or medicinally important genera including Cuminum, Daucus, Laser, Laserpitium and Thapsia among others. Recent molecular studies, based mostly on nrDNA ITS sequences, revealed that neither Daucus nor Laserpitium are monophyletic. To address issues of relationships and apply respective nomenclatural changes, we obtained additional ITS sequences as well as inde- pendent data from three plastid markers—rps16 intron, rpoC1 intron and rpoB-trnC intergenic spacer—for a comprehensive sample of the subtribe. We examined data for 260 accessions representing all genera of Daucinae and 81 of its ca. 93 species. Phylogenetic trees were estimated using maximum likelihood and Bayesian inference methods. The results indicate that former Laserpitieae constitute a paraphyletic grade at the base of the spiny-fruited members of Daucinae while traditionally delimited Daucus and Laserpitium are polyphyletic. To maintain a monophyletic Daucus, we suggest including the following genera and species into its synonymy: Agrocharis, Melanoselinum, Monizia, Pachyctenium, Pseudorlaya, Rouya, Tornabenea, Athamanta dellacellae and Cryptotaenia elegans. The species of Laserpitium occur in seven clades and only six species of the Laserpitium s.str. clade retain the generic name. Several species are transferred to Ekimia, Laser and Thapsia; additionally, a monospecific genus Siler is restored and a new genus, Silphiodaucus, is established. The inclusion of Ammodaucus into Thapsia suggested in an earlier study is not supported. The position of Laserpitium pseudomeum requires further study.
Background:
Four plastid regions, rpoB, rpoC1, matK, and trnH-psbA, have been recommended as DNA barcodes for plants. Their success in delimiting species boundaries depends on the existence of a clear-cut difference between inter- and intraspecific variability. We tested the ability of these regions to discriminate among closely related species in seven genera of flowering plants with different generation times (trees, perennials, and annuals). To ensure a maximum coverage of intraspecific diversity, and therefore to better evaluate the resolution power of each barcode, we applied a population genetics approach by sampling three to 45 individuals per species over a wide geographical range.
Results:
All possible combinations between loci were analysed, which showed that using more than one locus does not always improve the resolution power. The trnH-psbA locus was most effective at discriminating among closely related species (Acer, Lonicera, Geranium, and Veronica), singly or in combination. For Salix, Adenostyles, and Gentiana, the best results were obtained with the combination of matK, rpoB, and trnH-psbA. No barcoding gap was found within six genera analysed, excepting Lonicera. This is due to shared polymorphisms among species, combined with very divergent sequences within species. These genetic patterns reflect incomplete lineage sorting and hybridization events followed by chloroplast capture.
Conclusions:
Our results strongly suggest that adding trnH-psbA to the two obligate DNA barcodes proposed by the CBOL plant-working group (matK and rbcL) should be mandatory for closely related species. In our sampling, generation time had no influence on DNA barcoding success, as the best and worst identification successes were found for the two tree genera (Acer, 64 % success and Salix, 86 % failure). Evolutionary histories are the main factor influencing DNA barcoding success in the studied genera.
Basil is an important part of Lamiaceace family. Besides having various economical importance it also has diverse uses in the traditional medicines. It is rich in camphor, geraniol, linalool and linalyl acetate. The genus is highly variable and possesses wide range of diversity at the level of morphology, chemical and genetic make-up. For analyzing the variation existing in the genus Ocimum, the chemical analyses of its essential oils and DNA fingerprinting were prominently utilized. This is utmost helpful in plant improvement programs as well as developing a well-organized way to conserve the genetic wealth of the genus. In this paper, we have thrown light on how the combinatorial approach of studying the morphological traits, essential oil composition and DNA markers is useful in verifying the taxonomy of Ocimum. © 2016, International Journal of Pharmacognosy and Phytochemical Research. All rights reserved.
Pepper species exhibit broad genetic diversity, which enables their use in breeding programs. The objective of this study was to characterize the diversity between the parents of different species and their interspecific hybrids using morphological and molecular markers. The parents of Capsicum annuum (UFPB-01 and -137), C. baccatum (UFPB-72), and C. chinense (UFPB-128) and their interspecific hybrids (01x128, 72x128, and 137x128) were used for morphological and molecular characterization. Fruit length and seed yield per fruit (SYF) traits showed the highest variability, and three groups were formed based on these data. CVg/CVe ratio values (> 1.0) were calculated for leaf length (1.67) and SYF (5.34). The trait that most contributed to divergence was the largest fruit diameter (26.42%), and the trait that least contributed was pericarp thickness (0.33%), which was subject to being discarded. The 17 primers produced 58 polymorphic bands that enabled the estimation of genetic diversity between parents and hybrids, and these results confirmed the results of the morphological data analyses. The principal component analysis results also corroborated the morphological and random-amplified polymorphic DNA data, and three groups that contained the same individuals were identified. These results confirmed reports in the literature regarding the phylogenetic relationships of the species used as parents, which demonstrated that C. annuum was closer to C. chinense as compared to C. baccatum.
With ca. 200 species, the informally named Potato clade represents one of the larger subgroups of the estimated 1500 species of Solanum. Because its members include the potato (S. tuberosum), tomato (S. lycopersicum), and pepino (S. muricatum), it is the most economically important clade in the genus. These crop species and their close relatives have been the focus of intensive research, but relationships among major lineages of the Potato clade remain poorly understood. In this study, we use sequences from the nuclear ITS and waxy (GBSSI), and plastid trnT-trnF and trnS-trnG to estimate a phylogeny and further explore relationships within the Potato clade. With increased sampling over past studies, the Potato clade emerges as a strongly supported clade and comprises 12–13 subclades which, for the most part, correspond to traditionally recognized sections. Solanum sect. Regmandra is sister to the rest of the lineages of the Potato clade which are, in turn, organized into two major subclades: (1) sections Anarrhichomenum, Articulatum, Basarthrum, Etuberosum, Juglandifolia, Lycopersicoides, Lycopersicon, and Petota, and (2) sections Herpystichum and Pteroidea. As in all other studies including these groups, sections Etuberosum, Juglandifolia, Lycopersicoides, Lycopersicon, and Petota form a strongly supported clade. Solanum oxycoccoides, a high-elevation species endemic to north-central Peru, was tentatively assigned to several groups within Solanum based on morphological evidence, but instead the species represents an independent lineage within the Potato clade, sister to the first major subclade. A key to the sections of the Potato clade is provided.
Nucleotide sequences from the plastome are currently the main source for assessing taxonomic and phylogenetic relationships in flowering plants and their historical biogeography at all hierarchical levels. One major exception is the large and economically important genus
Quercus
(oaks). Whereas differentiation patterns of the nuclear genome are in agreement with morphology and the fossil record, diversity patterns in the plastome are at odds with established taxonomic and phylogenetic relationships. However, the extent and evolutionary implications of this incongruence has yet to be fully uncovered. The DNA sequence divergence of four Euro-Mediterranean Group Ilex oak species (
Quercus ilex
L.,
Q. coccifera
L.,
Q. aucheri
Jaub. & Spach.,
Q. alnifolia
Poech.) was explored at three chloroplast markers (rbcL, trnK/matK, trnH-psbA). Phylogenetic relationships were reconstructed including worldwide members of additional 55 species representing all
Quercus
subgeneric groups. Family and order sequence data were harvested from gene banks to better frame the observed divergence in larger taxonomic contexts. We found a strong geographic sorting in the focal group and the genus in general that is entirely decoupled from species boundaries. High plastid divergence in members of
Quercus
Group Ilex, including haplotypes shared with related, but long isolated oak lineages, point towards multiple geographic origins of this group of oaks. The results suggest that incomplete lineage sorting and repeated phases of asymmetrical introgression among ancestral lineages of Group Ilex and two other main Groups of Eurasian oaks (Cyclobalanopsis and Cerris) caused this complex pattern. Comparison with the current phylogenetic synthesis also suggests an initial high- versus mid-latitude biogeographic split within
Quercus
. High plastome plasticity of Group Ilex reflects geographic area disruptions, possibly linked with high tectonic activity of past and modern distribution ranges, that did not leave imprints in the nuclear genome of modern species and infrageneric lineages.
The substitution of low-cost or adulterated herbal products for high-priced herbs makes it important to be able to identify and trace herbal plant species and their processed products in the drug supply chain. PCR-based methods play an increasing role in monitoring the safety of herbal medicines by detecting adulteration. Recent studies have shown the potential of DNA barcoding combined with high resolution melting (Bar-HRM) analysis in herbal medicine identification. This method involves precisely monitoring the change in fluorescence caused by the release of an intercalating DNA dye from a DNA duplex as it is denatured by a gradual increase in temperature. Since the melting profile depends on the GC content, length, and strand complementarity of the amplification product, Bar-HRM analysis opens up the possibility of detecting single-base variants or species-specific differences in a short region of DNA. This review summarizes key factors affecting Bar-HRM analysis and describes how Bar-HRM is performed. We then discuss advances in Bar-HRM analysis of medicinal plant ingredients (herbal materia medica) as a contribution toward safe and effective herbal medicines.
American ginseng (derived from Panax quinquefolius) is one of the most widely used medicinal herbs in the world. Because of its high price and increasing demand, there are many adulterants on the market. The proposed internal transcribed spacer 2 (ITS2) has been used to identify raw medicinal materials, but it is not suitable for the identification of Chinese patent medicine ingredients. Therefore, a short barcode for the identification of processed American ginseng and its corresponding Chinese patent medicines would be profitable. In this study, 94 samples of American ginseng and Asian ginseng were collected from all over the world. The ITS2 region was sequenced, and a nucleotide signature was developed based on one single nucleotide polymorphism (SNP) site unique to American ginseng. The nucleotide signature (atcactcctt tgcgggagtc gaggcgg) consists of 27 bases over the length of the ITS2 sequence (420 bp). Furthermore, we also designed primer pairs to amplify the nucleotide signature; the specific primer pair 4F/4R has been found to be unique to the ginseng species and capable of amplifying the nucleotide signatures from Chinese patent medicines and decoctions. We used the nucleotide signature method to inspect ginseng products in Chinese patent medicines; 24 batches of Chinese patent medicine from stores in Beijing were amplified and sequenced successfully. Using the double peaks at the SNP sites of the nucleotide signature, 5 batches were found to be counterfeits, and 2 batches were found to contain adulterants. Thus, this nucleotide signature, with only 27 bp, has broadened the application of DNA barcoding in identification of decoctions, Chinese patent medicines and other ginseng products with degraded DNA. This method can rapidly identify ginseng products and could also be developed as an on-site detection method.
Phylogenetic relationships among 21 species in the Brassica genus, and Raphanus sativus in the family Brassicaceae, were estimated from sequences of the internal transcribed spacers, ITS1 and ITS2, in the nuclear DNA and the 5.8S rRNA gene, as well as from chloroplast DNA (cpDNA) sequences from the trnL intron, the trnL-F region, and chloroplast microsatellite (cpSSR) markers. The data from all three molecular datasets indicated that the “B genome” and “A/C genomes” of Brassica spp. originated via two different evolutionary pathways. Combining the nuclear and chloroplast sequences, Raphanus might be a hybrid between members of the “Nigra” and “Rapa/Oleracea” lineages, with the latter as the female parent. The chloroplast sequences and cpSSR data suggested that the cytoplasmic parents of the amphidiploid species B. carinata and B. juncea were B. nigra and B. rapa, respectively.
Taking into account the recommendations of the Codex Alimentarius Commission, the European Commission amended the European Food Labelling Directive 2000/13/EC by a list of ingredients to be labeled, irrespective of their amount in the final product.Therefore, suitable detection methods are needed in order to assure the surveillance of the established legislation. The simultaneous detection of DNA from different allergenic food ingredients by a ligation-dependent probe amplification (LPA) system is described.The approach applied in this study allows the detection of several targets in a onetube assay. The specific and sensitive LPA system for the detection of tree nuts (cashew, pecan nut, pistachio nut, hazelnut, macadamia nut, almond, walnut and brazil nut), peanut and sesame seeds was used for the analysis of commercially available samples.The method was shown to be suitable to detect allergenic ingredients and traces given in the voluntary specifications within the product liability in different food matrices of the retail samples.
Background and aims:
The origin of limes and lemons has been a source of conflicting taxonomic opinions. Biochemical studies, numerical taxonomy and recent molecular studies suggested that cultivated Citrus species result from interspecific hybridization between four basic taxa (C. reticulata, C. maxima, C. medica and C. micrantha). However, the origin of most lemons and limes remains controversial or unknown. The aim of this study was to perform extended analyses of the diversity, genetic structure and origin of limes and lemons.
Methods:
The study was based on 133 Citrus accessions. It combined maternal phylogeny studies based on mitochondrial and chloroplastic markers, and nuclear structure analysis based on the evaluation of ploidy level and the use of 123 markers, including 73 basic taxa diagnostic single nucleotide polymorphism (SNP) and indel markers.
Key results:
The lime and lemon horticultural group appears to be highly polymorphic, with diploid, triploid and tetraploid varieties, and to result from many independent reticulation events which defined the sub-groups. Maternal phylogeny involves four cytoplasmic types out of the six encountered in the Citrus genus. All lime and lemon accessions were highly heterozygous, with interspecific admixture of two, three and even the four ancestral taxa genomes. Molecular polymorphism between varieties of the same sub-group was very low.
Conclusions:
Citrus medica contributed to all limes and lemons and was the direct male parent for the main sub-groups in combination with C. micrantha or close papeda species (for C. aurata, C. excelsa, C. macrophylla and C. aurantifolia - 'Mexican' lime types of Tanaka's taxa), C. reticulata (for C. limonia, C. karna and C. jambhiri varieties of Tanaka's taxa, including popular citrus rootstocks such as 'Rangpur' lime, 'Volkamer' and 'Rough' lemons), C. aurantium (for C. limetta and C. limon - yellow lemon types - varieties of Tanaka's taxa) or the C. maxima × C. reticulata hybrid (for C. limettioides - 'Palestine sweet' lime types - and C. meyeri). Among triploid limes, C. latifolia accessions ('Tahiti' and 'Persian' lime types) result from the fertilization of a haploid ovule of C. limon by a diploid gamete of C. aurantifolia, while C. aurantifolia triploid accessions ('Tanepao' lime types and 'Madagascar' lemon) probably result from an interspecific backcross (a diploid ovule of C. aurantifolia fertilized by C. medica). As limes and lemons were vegetatively propagated (apomixis, horticultural practices) the intra-sub-group phenotypic diversity results from asexual variations.
Abstract— Commiphora (Burseraceae) is the most species-rich genus in the frankincense and myrrh family, whose widespread and often remote distribution in seasonally dry tropical forests globally has impeded efforts to resolve its taxonomy and investigate its systematic biology. Here we focus on establishing the origin and evolution of Commiphora species native to Madagascar, all of which are endemic. Recent work has uncovered 16 new Malagasy species, bringing the total number of island endemic species to 44 and comprising nearly 25% of the genus. Previous phylogenetic studies of Commiphora have indicated that Malagasy lineages immigrated to and radiated within the island twice. We test this biogeographic hypothesis more thoroughly using a nearly comprehensive sampling of species from Madagascar, an expanded sampling of Commiphora species native to other regions, and a greater depth of DNA sequence data from the nuclear (nrETS, nrITS) and chloroplast (psbA‐trnH, ndhF‐rpl32, and trnD‐trnT) genomes. Results from this expanded phylogenetic analysis include strong support for seven infrageneric clades within a monophyletic Commiphora, which we refer to as the ‘Lasiodisca,’ ‘Granulifera,’ ‘Saxicola,’ ‘Gariepensis,’ ‘Spinescens,’ ‘Arafy,’ and ‘Rhynchocarpa’ clades. Malagasy species comprise four distantly related lineages, including one species sister to all other Commiphora species (C. lasiodisca), the species-rich ‘Arafy,’ and ‘Rhynchocarpa’ clades, and one species embedded within the predominantly East African ‘Spinescens’ clade (C. simplicifolia). We describe the morphological and geographic affinities of each infrageneric and Malagasy lineage and identify priorities for future systematic study of the genus.
This study evaluated the feasibility of using the Ribulose Bisphosphate Carboxylase Large subunit gene (rbcL) and the Internal Transcribed Spacers 1 and 2 of the nuclear rDNA (nuITS1 and nuITS2) markers for identifying a very diverse, albeit poorly known group, of green microalgae from neotropical inland waters. Fifty-one freshwater green microalgae strains isolated from Brazil, the largest biodiversity reservoir in the neotropics, were submitted to DNA barcoding. Currently available universal primers for ITS1-5.8S-ITS2 region amplification were sufficient to successfully amplify and sequence 47 (92%) of the samples. On the other hand, new sets of primers had to be designed for rbcL, which allowed 96% of the samples to be sequenced. Thirty-five percent of the strains could be unambiguously identified to the species level based either on nuITS1 or nuITS2 sequences' using barcode gap calculations. nuITS2 Compensatory Base Change (CBC) and ITS1-5.8S-ITS2 region phylogenetic analysis, together with morphological inspection, confirmed the identification accuracy. In contrast, only 6% of the strains could be assigned to the correct species based solely on rbcL sequences. In conclusion, the data presented here indicates that either nuITS1 or nuITS2 are useful markers for DNA barcoding of freshwater green microalgae, with advantage for nuITS2 due to the larger availability of analytical tools and reference barcodes deposited at databases for this marker.
Legumes in the Omic Era provides a timely review of recent advances in legume genomics research and application. In this post-genomic era enormous amount of biological information is available which could be of huge potential use for crop improvement applications. This aspect of genomics assisted plant breeding is focused throughout the book for all the important grain legume crops. Role of functional genomics and importance of bioinformatics tools in present day genomics and molecular breeding research is also discussed in detail. Use of molecular tools for nutritional fortification of grain legume is briefly presented. A chapter also been contributed on fungal disease resistance to elucidate potential application of genomic tools in molecular breeding of grain legume species. The book contains fifteen chapters contributed by 50 scientists from different countries who are actively involved in analyzing and improving particular legume genome. This book will serve as reference resource to legumes researchers for use of genome information in improvement of major legume crops. © Springer Science+Business Media New York 2014. All rights are reserved.
The wide distribution of Valeriana officinalis as a herbal remedy as well as the considerably higher concentration of putative mutagenic valepotriate metabolites in other drug-delivering valerian species like Valeriana procera Kunth and Valeriana jatamansi Jones ex Roxb. illustrate the necessity of secure authentication of roots of Valeriana officinalis s.l., especially as the morphologically similar roots of the acutely toxic Veratrum album can be mistaken for those of Valeriana officinalis. We developed two DNA-based systems, a multiplex amplification refractory mutation system (MARMS), and a high-resolution melting curve analysis (HRMA) assay, both based on a sequence mutation within the atpB-rbcL region. With both methods, identification of Valeriana officinalis s.l. was possible. With the HRMA, the characteristic melting curve of 33 samples of Valeriana officinalis s.l. and of two commercial samples of Valerianae radix was distinct from the melting curves of all other Valeriana species (60 accessions), and from the closely related genera Centranthus and Valerianella. Since adulteration of Valeriana with toxic Veratrum species was reported previously, Veratrum primers were included in a multiplex PCR-HRM analysis. This system allowed the detection of a Veratrum admixture down to the level of 0.01 %. Although the advantages, in terms of sensitivity, specificity and practicality of the HRM for analysis of degraded plant material were superior to the MARMS assay, both methods are suitable for routine analysis. The results demonstrated the general ability of HRMA to detect specific (toxic) adulterations in drugs in a semiquantitative way.
The aim of this study was to delimit the taxa of the Crataegus rosei complex using an integrative approach that incorporates a suite of molecular (cpDNA and nuclear microsatellite markers), morphological, and geometric morphometric characters. One hundred and ten plants from 19 populations that encompass the entire distribution range of the species complex were collected and examined along with herbarium specimens. Parsimony and Bayesian inference analyses were run using morphological, molecular, and both the morphological and molecular data sets combined. Analyses to determine genetic structure based on microsatellite data and multivariate analyses incorporating geometric morphometrics were also done to identify differences in leaf shape. The results supported the recognition of two taxa: C. rosei with high levels of gene flow among its populations, remarkable morphological variation and a wide distribution range and C. rosei var. amoena, composed of a few isolated populations in the high elevation location of Cerro Potosí; a new specific epithet will be decided for the latter in accordance with the International Code of Nomenclature for algae, fungi, and plants.
A molecular phylogeny including c. 75% of all known species belonging to the closely related genera Anthemis and Cota (Compositae, Anthemideae) and based on nucleotide sequences from two plastid regions (psbA‐trnH and trnC‐petN spacers) and one nuclear marker (ITS) is presented. The molecular analyses were supplemented by a multivariate analysis of 25 micromorphological and anatomical characters. The results show incongruences among plastid and nuclear marker sets, which indicate that hybridisation may have played an important role in the evolution of this morphologically diverse plant group. However, molecular and morphological data both support a sister‐group relationship between four perennial species (A. calcarea, A. fruticulosa, A. marschalliana, A. trotzkiana) and the main clades of Cota and Anthemis. We propose segregation of these from Anthemis as a new genus, Archanthemis. With the exclusion of these four taxa, both the nrDNA and the cpDNA datasets support the monophyly of Anthemis. Further molecular and morphological evidence is provided for the generic independence of the genus Cota. The traditional infrageneric classifications of Anthemis and Cota are not entirely supported by the phylogenetic analyses as a picture of intense reticulate evolution among the different lineages emerges.
Generic delimitation of Cyathocalyx and Drepananthus has been controversial, with some authors recognizing them as distinct genera, and others recognizing a more broadly defined Cyathocalyx, inclusive of Drepananthus. Some doubt also exists regarding the relationships between these taxa and Cananga. Molecular phylogenetic analyses are presented based on combined psbA‐trnH spacer, trnL‐F, matK and rbcL sequences. Results indicate that Cananga, Cyathocalyx s.str. and Drepananthus form three generally well‐supported clades, although with inadequate resolution of relationships among the three clades. Morphological variation is re‐evaluated, and the narrower delimitation of Cyathocalyx proposed, necessitating 21 new nomenclatural combinations following the recognition of Drepananthus as a distinct genus. Divergence times are estimated using an uncorrelated lognormal distributed (UCLD) relaxed molecular clock. Historical biogeographical analysis suggests that the ambavioid lineage originated in Africa, with subsequent dispersal into Asia. Alternative hypotheses for this dispersal, involving rafting on the Indian tectonic plate versus migration via the extensive boreotropical forests associated with the Eocene thermal maximum, are evaluated, and the latter route identified as the most consistent with the divergence age estimates and the geological and palaeoclimatic data.
The banana family (Musaceae s.str.; Zingiberales), an economically important tropical group of plants, includes three genera, Musa, Ensete and Musella, and possibly 41 species. We performed phylogenetic analyses of a total of 39 accessions covering 28 species in the Musaceae and five outgroup species using nuclear ribosomal ITS and chloroplast trnL‐F sequences. Outgroups were chosen from the closely related families Lowiaceae, Strelitziaceae, and Heliconiaceae. Our results suggest that Musaceae s.str. is monophyletic. Three main internal clades are well‐supported within the family. The genus Musa is comprised of two of these clades, and Musella plus Ensete make‐up the third clade. The sectional classification system of Musa based on chromosome numbers is not supported by DNA sequence evidence. Both inflorescence orientation (erect or pendent) and chromosomal number in Musa, which were characters traditionally thought to be diagnostic in sectional classification, are homoplasious traits in the family. The disjunct distribution of living members of the genus Ensete in tropical Asia and Africa with a fossil species described from the Eocene of Oregon in North America may be an example of the distributional retreat of the Boreal Tropics. The phylogenetic position of the monospecific Musella as sister to the African clade of Ensete suggests that the single species in this lineage is a highly specialized member not warranting generic status. Evidence from the molecular phylogenetic investigations highlights the evolutionary diversification and biogeographic context of this plant group, and suggests additional taxonomic investigations of both Musa and Ensete are in order.
We investigated the species and genome relationships among 82 Avena accessions representing thirteen diploid species (A and C genomes), six tetraploids (AB and AC genomes) and four hexaploids (ACD genome) to infer the evolutionary pathways in Avena using the plastid matK gene and the trnL‐F region. The matK and trnL‐F sequences pointed to the A‐genome diploid species as maternal parents of the polyploid species. Furthermore, different A‐genome diploid species might have served as the A genome donor of several different polyploid species. The probable ancestor of most hexaploid species, of the AC‐genome tetraploids, and of the AB‐genome species A. agadiriana was A. wiestii (As genome). The likely donor of the other three AB‐ genome tetraploids was A. hirtula (also As genome) and A. damascena (Ad genome) appears to be the A genome donor of the hexaploid A. fatua. The A genome origin of A. fatua differs from that for the other hexaploid Avena species, which is different from the common assumption that the hexaploid species evolved from a single hexaploid ancestor followed by gain or loss of domestication genes. Thus, several separate maternal lineages might be involved in different polyploid species.
Mentheae are the largest tribe in the family Lamiaceae and economically important, including herbs like mint, sage and thyme. The evolutionary history of this tribe was reconstructed based on ITS and trnL‐trnF spacer sequence data of 71 species, representing 47 out of 65 genera. The resulting phylogeny was used to analyse the distribution of selected morphological characteristics such as sexine ornamentation of pollen, nutlet shape with existence of abscission scar and its form, and trichome types. Two monophyletic groups are recognized, which largely correspond to the current subtribal circumscription. Subtribe Salviinae is monophyletic, including the genus Melissa which was a genus of uncertain affinity in Mentheae. Subtribe Menthinae is not monophyletic since Cleonia, Horminum, Hyssopus, Lycopus and Prunella are more closely related with subtribe Nepetinae. Although we could not detect any morphological synapomorphies for each clade, morphological variation seems to be correlated with the molecular phylogeny. A circular abscission scar without distinct lateral areole occurred mainly in Salviinae, while the majority of the species of Mentheae and Nepetinae had a clear areole at the abscission scar. In addition, a reticulate sexine ornamentation is rather common in the Menthinae clade.
Ononis L. (Fabaceae, Papilionoideae) is a common genus in the circum‐Mediterranean region. To infer phylogenetic relationships, 69 Ononis species were analyzed using plastid trnL‐F and nuclear ITS DNA sequences. Trees resulting from maximum parsimony analysis and Bayesian inference provide evidence that Ononis is monophyletic but contradict the traditional subgeneric division in O. sects. Ononis and Natrix. However, many of the 22 subsections of the genus are well supported by the molecular data. Phylogenetic reconstructions indicate five major lineages within the genus, which are morphologically supported by peduncle length and flower color, characters which also confirm the polyphyly suggested for O. subsect. Reclinatae. Ononis subsects. Antiquae and Rhodanthae form a basally branching monophylum, confirming the findings of previous studies. An examination of the life strategies dominant in different clades and the climatic conditions in the habitats of the species in the light of the molecular data suggests that O. subsects. Ononis, Mauritanicae and Canariensis are secondarily perennial, with the latter two serving as examples of high altitude woodiness and insular woodiness.
Although extensive studies have demonstrated that several genera formerly placed in the family Flacourtiaceae are the closest relatives of Populus-Salix. However, the closest relative or relatives remain uncertain due to the relatively low phylogenetic resolution and insufficient phylogenetic information. In addition, the question as to whether Populus and Salix are truly monophyletic remains open because of the absence of comprehensive studies with large-scale samples of both genera. In the present study, 54 species representing 11 genera of Salicaceae sensu lato (now including the genera of Flacourtiaceae) were studied using a combined rbcL and matK gene sequence dataset with a total length of 2134 bp, to explore the closest genera that are relatives of Populus-Salix and to determine the relationships between Populus and Salix. The resulting phylogenetic trees showed high resolution in each of the main clades. The results confirmed that (1) Idesia and Bennettiodendron are the closest relatives to Populus-Salix, followed by Poliothyrsis, and (2) Populus and Salix are a truly monophyletic lineage in which the most recent ancestor is shared by the two genera. This finding is significant for a wide variety of evolutionary studies and for understanding the classification and biosystematics of Salicaceae.
The genus Citrus (Aurantioideae, Rutaceae) is the most important fruit crops all around the world. As many cultivated and weedy species/varieties exist in this genus, botanists hold different opinions on the taxonomy and phylogeny of the genus Citrus all the time. To evaluate the taxonomy and phylogenetic relationship of Citrus species, we collected 22 Citrus species and evaluated the genetic diversity and phylogeny based on the genetic variation of the ribosomal RNA (rRNA) 5S gene sequences. The 5S DNA alignment showed high nucleotide variations and sequence length difference among investigated species. In the rRNA 5S gene, four nucleotide sites were found to be easily nucleotide-substituted and hybridization-reflective. Some Citrus species, including C. kinokuni, C. unshiu, C. sinensis, C. leiocarpa, C. tagerina andC. hybrid are found to be hybridization-susceptible species. The analysis result of genetic distance suggested that C. clemntina, C. hybrid, C. leiocarpa, C. nippokoreana, C. kinokuni and C. tagerina showed low values with each other and divided into one group in the rRNA 5S phylogenetic tree. Others were grouped together in the 77% similarity criterion. According to six-tribe discrimination method, the clading seen from the 5S phylogenetic tree suggested that the species belonging to Sinocitrus tribe showed higher sequence variation than other tribes. This work provides more sequence evidences for Citrus species identification and helps us further understand the phylogenetic relationships and species identification of the genus Citrus. The results would help citrus breeding and germplasm conservation programs.
Crocus sativus L. and Aloe barbadensis Mill. are well known functional ingredients in health and food products and are known for their medicinal properties as antioxidant, antidepressant, relaxant and cathartic agents. In this study, reliable quality control markers were developed for the quality assurance of C. sativus and A. barbadensis at raw material source as well as in finished herbal products. DNA based sequence-characterized amplified region (SCAR) markers were developed from randomly amplified polymorphic DNA (RAPD) markers specific for both the species to detect its adulteration in commercial products. Developed RAPD markers were cloned and sequenced and SCAR primers were generated SCAR markers upon PCR amplification with genomic DNA. These specific SCAR markers enabled unequivocal detection of adulterants in the commercially procured polyherbal medicine. C. sativus could not be identified through SCAR markers, instead SCAR amplicon of Carthamus tinctorius was detected suggesting that although being labelled as one of the constituents, C. sativus may not have been used to prepare the polyherbal medicine. This simple analytical strategy could be used for large scale screening of medicinal plants at raw material source as well as in finished polyherbal medicine.
Objective: DNA molecular identification of medicinal plants in Angelica L. was carried out based on the internal transcribed spacer (ITS) of ribosomesequence variation. Methods: One hundred and fifty-eight individuals of 32 species from Angelica L. were selected, and ITS sequences were used to evaluate the differences among these species. Obtained sequences were analyzed using MEGA6.0 program with the kimura-2-parameter (K2P) model. Meanwhile, phylogenetic trees were constructed, included the neighbor-joining (NJ) tree, the maximum-likelihood (ML) tree, and the maximum-parsimony (MP) tree, and we determined the identification ability of ITS sequences and the phylogenetic relationships among the species in Angelica L. Results: The minimum interspecific distance is far greater than the maximum intraspecific distance in ITS region. The phylogenetic analysis showed that the species in Angelica L. can be easily differentiated according to their strong bootstrap support and relationships. Conclusion: ITS region as DNA barcode can distinguish the medicinal plants in Angelica L. and would be a foundation for the aid of molecular identification of the species in Angelica L. © 2016, Editorial Office of Chinese Traditional and Herbal Drugs. All right reserved.
Objective: To identify the authenticity of Bupleuri Radix in the market of Hebei province. Methods: In this research, the ITS2 sequence of 28 samples of Bupleuri Radix that were bought from the market in Hebei province was amplified by PCR, and the sequence of seven samples was downloaded from GenBank. Aligner CodonCode software was used for sequence alignment, the MEGA6.0 analysis was used to calculate the K2P distance, and the mutation sites of each sequence was analyzed. Results: Among the 28 Bupleuri Radix samples, there were 21 samples for Bupleurum chinese, 3 for B.smithii, and 4 for the false. Conclusion: ITS2 sequence can be used as an effective method to identify the authenticity of Bupleuri Radix, it is great value to the medicine market standard and the clinical medication safety. © 2016, Editorial Office of Chinese Traditional and Herbal Drugs. All right reserved.
Objective: To evaluate the suitable candidate DNA barcoding of plants in Rehmannia Libosch. ex Fisch. et Mey., and unravel the origin of cultivated R. glutinosa. Methods: Nuclear DNA ITS and chloroplast gene psbA-trnH, rbcL, and matK sequences of species in Rehmannia Libosch. ex Fisch. et Mey. were amplified and sequenced. The Kimura 2-Parameter (K2P) distances were calculated. Identification analyses were performed using Nearest Distance, BLAST1, and Neighbor-Joining (NJ) methods. Results: A comparison on the sequences within species in Rehmannia Libosch. ex Fisch. et Mey. indicated that the rbcL sequences were identical, the matK sequences were similar by 99.9%-100%. All inter-specific distances (ITS and psbA-trnH data) were far higher than all intra-specific genetic distances. Minimum interspecific distance (ITS and psbA-trnH data) were higher than coalescent depth. The NJ trees (ITS, psbA-trnH, and combined data) indicated that all population of R. glutinosa formed a monophyletic clade [Bootstrap (BS)=96%, 55%, and 58%]. The clades including R. glutinosa and R. solanifolia were clustered with R. piasezkii and R. elata in ITS and combined trees (BS=55% and 68%), and clustered with R. piasezkii and R. chingii in psbA-trnH tree (BS=80%). The NJ trees (ITS, psbA-trnH, and combined data) supported that three cultivated varieties of R. glutinosa were clustered with wild populations from Wenxian, Zhengzhou, Nanyang, and Beijing (BS=51% and 69%). Conclusion: Chloroplast genes rbcL and matK can not be used to identify the medicinal plants of Rehmannia Libosch. ex Fisch. et Mey. ITS and psbA-trnH are two efficient barcodes for authentication of R. glutinosa and its relative species. R. piasezkii and R. chingii may be as both parental species of tetraploid R. glutinosa. Furthermore, it appears that native wild populations are involved in the origin of cultivated R. glutinosa in Wenxian county. © 2016, Editorial Office of Chinese Traditional and Herbal Drugs. All right reserved.
Sugarcane (Saccharum hybrid cultivar) ranks among the world's top 10 food crops and annually provides 60–70% of the sugar produced worldwide. Despite its economic importance there has been no large-scale systematics study of genus Saccharum and the existing model of sugarcane origins has remained largely unchallenged for almost 50 years. For the first time, we have assembled the complete plastid genomes of Miscanthus floridulus (first report for this genus), Saccharum spontaneum and Saccharum officinarum allowing us to elucidate the phylogenetic origins of Saccharum s.s. species. We demonstrate that Saccharum s.s. is divided into four species, with S. spontaneum diverging from the remainder of the genus about 1.5 million years ago and S. robustum diverging 750,000 years ago. Two separate lineages, one leading to S. officinarum and the other leading to modern hybrid cultivars diverged from S. robustum 640,000 years ago. These findings overturn all previous hypotheses on sugarcane origins, demonstrating that sugarcane's antecedents could not have arisen by human action. All modern cultivars share a common Polynesian origin, whereas Old World canes, S. barberi and S. sinense, cluster as a distinct S. officinarum lineage. This makes modern cultivars a distinct species of genus Saccharum, and we formally propose the name Saccharum cultum for the ancestor of all lineages currently classified as Saccharum hybrid cultivars.
Phellodendri Cortex is derived from the dried barks of Phellodendron genus species, has been extensively used in traditional Chinese medicine. The cortex is divided into two odorless crude drugs Guanhuangbo and Huangbo. Historically, it has been difficult to distinguish their identities due to a lack of identification methods. This study was executed to confirm the identity and to ensure the species traceability of Phellodendri Cortex. In the current study, ananalys is based on the ITS (internal transcribed spacer) and psbA-trnH intergenic spacer (psbA-trnH) barcodes and HPLC (high-performance liquid chromatographic) fingerprinting was carried out to guarantee the species traceability of Guanhuangbo and Huangbo. DNA barcoding data successfully identified the three plants of the Phellodendron genus species by ITS+psbA-trnH, with the ability to distinguish the species origin of Huangbo. Moreover, the psbA-trnH data distinguished Guanhuangbo and Huangbo except to trace species. The HPLC fingerprint data showed that Guanhuangbo was clearly different from Huangbo, but there was no difference between the two origins of Huangbo. Additionally, the result of HCA (hierarchical clustering analysis), based on chlorogenic acid, phellodendrine, magnoflorine, jatrorrhizine, palmatine and berberine, was consistent with the HPLC fingerprint analysis. These results show that DNA barcoding and HPLC fingerprinting can discriminate Guanhuangbo and Huangbo. However, DNA barcoding is more powerful than HPLC fingerprinting for species traceability in the identification of related species that are genetically similar. DNA barcoding is a useful scientific tool to accurately confirm the identities of medicinal materials from multiple sources.
• Key messageWe show that joint multivariate analyses of leaf morphological characters and molecular-genetic markers improve the taxonomic assignment in hybridizing European white oaks. However, model-based approaches using genetic data alone represent straightforward alternatives to laborious, detailed morphological assessments. • ContextIn European white oaks, species delimitation is debated because of large overlap of morphological characteristics likely due to hybridization. • AimsWe tested whether joint multivariate analyses of leaf morphology and molecular markers improve the identification of three oak species (Quercus petraea, Quercus pubescens, Quercus robur) compared to approaches using morphological or genetic variables only. • Methods
We assessed 13 leaf morphological characters and applied eight nuclear microsatellite markers in almost 1400 trees of 71 oak populations across Switzerland. We performed two multivariate approaches with three variable sets (morphology, genetics, combined) and assessed their performance in separating the taxa. We also compared the taxon assignment to a model-based clustering approach (Structure) based on genetic data alone. • ResultsA joint use of morphological and genetic variables led to an improved taxon assignment. Whereas Q. robur could clearly be separated from the two other taxa, there was a certain overlap between Q. petraea and Q. pubescens. The Structure clustering led to the same taxon assignment in 85 % of the individuals. • Conclusion
It is important to consider both morphological and genetic properties in morphologically similar and hybridizing species. However, it might be more efficient to concentrate only on genetic markers than on time-consuming morphological assessments.
Gelidioid specimens from Singapore were analyzed using sequences of cox1 and rbcL to identify the species. Both datasets revealed the presence of four species: Gelidiella acerosa, Pterocladiella bartlettii, Pterocladiella caerulescens and Gelidium sentosaense sp. nov., described here. Gelidium sentosaense formed small tufts up to 1 cm high and occurred in monospecific patches on cement beds at the outlet of drainpipes at intertidal areas. A morphological investigation of fresh specimens collected from Singapore revealed that G. sentosaense was similar to Gelidium pusillum, which is limited to North Atlantic coasts. However, it was distinguished by longitudinal striations present on the surface of branches and rhizines scattered in the inner cortex and medulla. Phylogenetic analyses of rbcL and cox1 sequences revealed that G. sentosaense consistently formed a clade with Gelidium sp. from Oman and Gelidium pluma from Hawaii but it was distinct from previously described Gelidium species.
Single-copy nuclear genes (SCNGs) are effective tools for phylogenetic analysis in plants. In this study, genetic variation and interspecific relationships in the genus Rehmannia were investigated by a novel set of SCNG markers. In total, 12 pairs of primers were successfully amplified and sequenced in six Rehmannia species after screening 258 candidate genes from the transcriptome datasets of Rehmannia glutinosa. The moderate to high level of nucleotide polymorphisms were identified in these SCNG markers, and nucleotide diversity (π) per locus ranged from 0.00498 to 0.01100, with 6-10 haplotypes. R. chingii and R. piasezkii exhibited a high level of genetic variation, while R. glutinosa and R. solanifolia had no genetic diversity. Phylogenetic analyses suggested that R. glutinosa - R. solanifolia, R. elata - R. piasezkii should be classified as one species, respectively. Molecular dating supported the rapid speciation of Rehmannia in Pleistocene. The single-copy DNA markers developed here would be also helpful for the study on the process of speciation and the population dynamic history of Rehmannia.
Brassica species have an economic and medicinal importance. Estimation of the amount and distribution of genetic diversity within Brassica species is essential for establishing efficient management, conservation and breeding practices. This review discusses the taxonomy, gene pool, and Brassica-derived phytochemicals and their nutraceutical importance. It also surveys the recently advanced studies of the genetic diversity and phylogenetic studies of Brassica species at the level of morphological, cytological, biochemical and molecular markers that have proven to be useful for evaluating the genetic variation, taxonomic relationships and species identity, and could be useful for improving Brassica crops through future promising breeding programmes.
OBJECTIVE: To discuss the taxonomic status of Pueraria lobata and its sibling species DC. in China, and to provide DNA molecular markers for authenticating the Chinese crude drug Gegen. METHODS: The internal transcribed spacers (ITS) including 5.8S rDNA, and partial 18S rDNA, 26S rDNA of 3 sibling species of Pueraria DC. in China were determined by using direct sequencing of PCR product. RESULTS: In DNA DIST analysis, the range of diversity between Pueraria lobata and P. thomsonii was 2.10% based on ITS1 and 2.09% based on ITS2. The range of diversity among the P. montana, P. lobata and P. thomsonii was above 8.14% based on ITS1 and 12.82% based on ITS2. Phylogram tree based on ITS and 5.8S sequence data indicated that P. lobata and P. thomsonii was closely related, while the genetic distance between P. montana and P. lobata was long. CONCLUSION: Base on the molecular characters, P. thomsonii may be taken as variant of P. lobata, but P. montana is independent species. The ITS sequence is good molecular marker for identification of the Chinese medicine Gegen.
Objective: To screen the DNA barcodes for identifying the species in genus Aesculus Linn. Methods: The PCR amplification and sequencing efficacy, intraspecific and interspecific variation, and identification efficiency of nuclear ITS, ITS2, chlroplast psbA-trnH, rbcL, and matK sequences from 42 samples of 10 species in genus Aesculus Linn. were investigated. The screened sequences were determined by barcoding gap and NJ tree clustering analysis. Results: The PCR amplification and sequencing efficacy of psbA-trnH were 100%, the interspecific minimum variation was larger than the intraspecific maximum variation, and psbA-trnH had the highest identification efficacy. The psbA-trnH sequence had less polymerization of intraspecific and interspecific variation. By constructing the NJ tree, thorough verifying the three original plants could be used to separate with other related species. Conclusion: The psbA-trnH sequence is a powerful, though not perfect, barcode for the identification of species in Aesculus Linn.
In order to reexamine the classification of Japanese Perilla plants, the phylogenetic relationships were investigated by RFLP (restriction fragment length polymorphism) and RAPD (randomly amplified polymorphic DNA) analyses using total DNAs. The results firmly supported the revised taxonomical classification on the basis of the morphological characteristics and cytogenetic experiments, and also indicated that P. citriodora was the most closely related species to the cultivated species, P. frutescens, among wild species.
Actinidia, a genus of dioecious vines, contains several species that bear fruit of economic value, among them the familiar kiwifruit. We have applied a set of nine microsatellite markers to leaf material of three Actinidia species, at three different ploidy levels. At each level parental and sibling plants were used to test the ability of the marker set to distinguish particular genotypes from closely related material. Eight of the nine markers, capable of distinguishing cultivars related to Actinidia deliciosa cv. Hayward, hexaploid, were polymorphic in an A. deliciosa family, and eight were also polymorphic in a family of the closely related diploid species Actinidia chinensis. In the more distantly related Actinidia arguta, only four of the nine markers were informative. In the three species themarker kit did distinguish individual genotypes in each family.