Complete chloroplast genomes of eight Delphinium taxa (Ranunculaceae) endemic to Xinjiang, China: insights into genome structure, comparative analysis, and phylogenetic relationships.

BACKGROUND: Delphinium L. represents a taxonomically intricate genus of significant phylogenetic and economic importance in Ranunculaceae. Despite the existence of few chloroplast genome datasets, a comprehensive understanding of genome structures and selective pressures within the genus remains unknown. Furthermore, several taxa in this genus are exclusively found in Xinjiang, China, a region renowned for its distribution and diversity of Chinese and Central Asian Delphinium species. Therefore, investigating the features of chloroplast genomes in this area will provide valuable insights into the evolutionary processes and phylogenetic relationships of the genus. RESULTS: In this study, the eight newly completed chloroplast genomes are examined, ranging in length from 153,979 bp to 154,284 bp. Alongside these, analysing six previously reported taxa re-annotated in Delphinium, 111 unique genes are identified across all samples. Genome structure, distributions of simple sequence repeats and short dispersed repeats, as well as gene content are similar among these Delphinium taxa. Nine hypervariable intergenic spacers and protein coding regions, including ndhF-trnL(TAG), rpl16-intron, rpl33, rps15, rps18, trnK(TTT)-trnQ(TTG), trnP(TGG)-psaJ, trnT(GGT)-psbD and ycf1, are identified among 13 perennial Delphinium. Selective pressure and codon usage bias of all the plastid genes are performed within 14 Delphinium taxa. Phylogenetic analysis based on 14 Delphinium plastomes, alongside two Aconitum (Ranunculaceae) species serving as outgroup taxa, reveals the monophyletic nature of Delphinium. Our findings further discern Delphinium into two distinct clades: perennial species (clade I) and annual species (clade II). In addition, compared with the nrDNA ITS topology, cytological data and morphological characters, D. mollifolium and D. maackianum showed potential involvement in hybridization or polyploidization processes. Excluding these two species, the perennial Delphinium (clade I) exhibits a stronger consistency with the morphology-based system that utilized seed morphology. CONCLUSION: This study represents the first comprehensive analysis of plastomic variations among Delphinium taxa, based on the examination of 14 complete plastomes. The chloroplast genome structure of Delphinium is similar to other angiosperms and possesses the typical quadripartite structure with the conserved genome arrangement and gene features. In addition, the variation of non-coding regions is larger than coding regions of the chloroplast genome. Through DNA sequence divergence across Delphinium plastomes and subsequent phylogenomic analyses ndhF-trnL(TAG) and ycf1 are identified as promising molecular markers. These highly variable loci held significant potential for future phylogenetic and phylogeographic studies on Delphinium. Our phylogenomic analyses based on the whole plastomes, concatenation of 132 unique intergenic spacer regions, concatenation of 77 unique protein-coding genes and nrDNA ITS, all support the monophyly of Delphinium and perennial taxa clusters together into one clade within this genus. These findings provide crucial data for systematic, phylogenomic and evolutionary research in the genus for future studies.

Chloroplast genomic data provide new and robust insights into the phylogeny and evolution of the Ranunculaceae.

The family Ranunculaceae, a member of early-diverging eudicots that is increasingly being used as a model for the study of plant developmental evolution, has been the focus of systematic studies for centuries. Recent studies showed that the family can be divided into 14 tribes, with Glaucideae, Hydrastideae, and Coptideae being the successive basal-most lineages. The relationships among the remaining 11 tribes, however, remain controversial, so that a clear picture of character evolution within the family is still lacking. In this study, by sequencing, assembling and analyzing the chloroplast (cp) genomes of 35 species representing 31 genera of the 14 tribes, we resolved the relationships among the tribes and genera of the Ranunculaceae and clarified several long-standing controversies. We found that many of the characters that were once widely used for taxonomic and systematic considerations were actually results of parallel, convergent or even reversal evolution, suggestive of unreliability. We also found that the family has likely experienced two waves of radiative evolution, through which most of the extant tribes and genera were generated. Notably, both waves of radiation were correlated with the increase in the temperature of the earth, suggesting that global warming may have been the driving force of the radiation events. Based on these observations, we hypothesize that global warming and the associated decrease in the type and number of animal pollinators may have been the main reason why taxa with highly elaborate petals as well as those without petal were generated during each of the two waves of radiation.

[New records of medicinal plants in Tibet].

By the fourth survey of Chinese medicinal resources, new medicinal plants records of 2 genera and 5 species were reported in Tibet. They are two genera Rhynchoglossum and Asteropyrum, and five species including Rh. obliquum, A. peltatum, Urena repanda, Schefflera khasiana and Mimulus tenellus. All the voucher specimens are preserved in Herbarium of Tibet Agriculture and Animal Husbandry University.

Comparative Analysis of the Chloroplast Genomes of the Chinese Endemic Genus Urophysa and Their Contribution to Chloroplast Phylogeny and Adaptive Evolution.

Urophysa is a Chinese endemic genus comprising two species, Urophysa rockii and Urophysa henryi. In this study, we sequenced the complete chloroplast (cp) genomes of these two species and of their relative Semiquilegia adoxoides. Illumina sequencing technology was used to compare sequences, elucidate the intra- and interspecies variations, and infer the phylogeny relationship with other Ranunculaceae family species. A typical quadripartite structure was detected, with a genome size from 158,473 to 158,512 bp, consisting of a pair of inverted repeats separated by a small single-copy region and a large single-copy region. We analyzed the nucleotide diversity and repeated sequences components and conducted a positive selection analysis by the codon-based substitution on single-copy coding sequence (CDS). Seven regions were found to possess relatively high nucleotide diversity, and numerous variable repeats and simple sequence repeats (SSR) markers were detected. Six single-copy genes (atpA, rpl20, psaA, atpB, ndhI, and rbcL) resulted to have high posterior probabilities of codon sites in the positive selection analysis, which means that the six genes may be under a great selection pressure. The visualization results of the six genes showed that the amino acid properties across each column of all species are variable in different genera. All these regions with high nucleotide diversity, abundant repeats, and under positive selection will provide potential plastid markers for further taxonomic, phylogenetic, and population genetics studies in Urophysa and its relatives. Phylogenetic analyses based on the 79 single-copy genes, the whole complete genome sequences, and all CDS sequences showed same topologies with high support, and U. rockii was closely clustered with U. henryi within the Urophysa genus, with S. adoxoides as their closest relative. Therefore, the complete cp genomes in Urophysa species provide interesting insights and valuable information that can be used to identify related species and reconstruct their phylogeny.

A molecular phylogeny of Dichocarpum (Ranunculaceae): Implications for eastern Asian biogeography.

East Asia is characterized by high levels of species diversity and endemism. However, the biogeographical patterns and processes underlying the distribution of biodiversity within the area are still poorly known. In this study, we used plastid (matK, trnL-F, and trnH-psbA) and nuclear (ITS) DNA sequences to investigate the historical biogeography of Dichocarpum (Ranunculaceae), an eastern Asian endemic genus throughout warm-temperate and subtropical forests of the area. Phylogenetic analyses strongly support Dichocarpum as monophyletic, which contains two major clades. Clade I corresponds to section Hutchinsonia, and clade II includes sections Dichocarpum and Fargesia. Section Dichocarpum and its subsections Dalzielia and Dichocarpum are not recognized as monophyletic. Our results suggest that the most recent common ancestor of Dichocarpum occurred in central China and Japan in the earliest Early Miocene, and thus support an ancient vicariance event between Japan and China. Within mainland China, three migrations at the species level were hypothesized to explain the expansion of Dichocarpum from central China to southeastern Yunnan, Hengduan mountains, and eastern Himalaya. These migration events occurred in the Late Miocene to Early Pliocene, which may be associated with the uplift of the southeastern Qinghai-Tibetan Plateau and accordingly the expansion of subtropical forests in China around that period. A migration or dispersal from central China to Taiwan was inferred in the Early Pleistocene, which supports the close floristic affinity between Taiwan and mainland China. This study contributes to our knowledge on the historical biogeography of plants in eastern Asia.

Cytogeography of the subalpine marsh marigold polyploid complex (Caltha leptosepala s.l., Ranunculaceae).

PREMISE OF THE STUDY: Unrecognized variation in ploidy level can lead to an underestimation of species richness and a misleading delineation of geographic range. Caltha leptosepala (Ranunculaceae) comprises a complex of hexaploids (6x), rare nonaploids (9x), and dodecaploids (12x), all with unknown distributions. We delineate the geographic distribution and contact zones of the cytotypes, investigate morphologies of cytotypes and subspecies, and discuss the biogeography and evolutionary history of the polyploid complex. METHODS: Using cytologically determined specimens as reference, propidium iodide flow cytometry was performed on silica-dried samples and herbarium specimens from across the range of C. leptosepala s.l. Genome size estimates from flow cytometry were used to infer cytotypes. A key morphological character, leaf length-to-width ratio, was measured to evaluate whether these dimensions are informative for taxon and/or cytotype delimitation. KEY RESULTS: Dodecaploids were more northerly in distribution than hexaploids, and a single midlatitude population in the Northern Rockies yielded nonaploids. Genome size estimates were significantly different between all cytotypes and between hexaploid subspecies. Leaf length-to-width ratios were significantly different between subspecies and some cytotypes. CONCLUSIONS: Caltha leptosepala presents clear patterns of cytotype distribution at the large scale. Marked differences in morphology, range, and genome size were detected between the hexaploid subspecies, C. leptosepala subsp. howellii in the Cascade-Sierra axis and C. leptosepala subsp. leptosepala in the Rockies. Sympatry between cytotypes in the Cascades and a parapatric distribution in the Northern Rockies suggest unique origins and separate lineages in the respective contact zones.

[Identification of radix et rhizoma clematidis and its adulterants using DNA barcoding].

This study provides the candidate sequences in the identification of Radix et Rhizoma Clematidis and its adulterants using DNA barcoding. We amplified and sequenced the region psbA-trnH, with the data of 284 sequences from GenBank, the differential intra- and inter-specific divergences, genetic distance, barcoding gap were used to evaluate five barcodes, and the identification efficiency was assessed using BLAST1 and Nearest Distance methods. The results showed that psbA-trnH barcodes performed high identification efficiency and inter-specific divergences among the five different DNA barcodes. Analysis of the barcoding gap and NJ tree showed psbA-trnH was superior to other barcodes. Based on the identification and PCR amplification efficiency, psbA-trnH can be the ideal barcode to identify Radix et Rhizoma Clematidis and its adulterants accurately.

A phylogeny of Delphinieae (Ranunculaceae) shows that Aconitum is nested within Delphinium and that Late Miocene transitions to long life cycles in the Himalayas and Southwest China coincide with bursts in diversification.

The tribe Delphinieae (Ranunculaceae) comprises two species-rich genera, Aconitum and Delphinium, the latter including Consolida and Aconitella. The 650-700 species are distributed in Eurasia and North America; three species occur on tropical African mountains. Maximum likelihood analyses of 2088 aligned nucleotides of plastid and nuclear sequences obtained from up to 185 species of Delphinieae from throughout the geographic range (plus relevant outgroups) show that three short-lived (facultative annual or biennial) Mediterranean species belonging to Delphinium subgenus Staphisagria are the sister clade to all other Delphinieae, implying that Staphisagria needs to be raised to genus status if Delphinium and Aconitum are to become mutually monophyletic. Molecular clock dating suggests an origin of the sampled Delphinieae in the Early Oligocene (c. 32.3 Ma) and expansion to North America of Aconitum and Delphinium around 3.3 and 2.9 Ma ago, respectively; the East African Mts. were reached by long-distance dispersal some 2.4 Ma ago, coincident with the major uplift of the East African Rift system. The ancestral growth form of the Delphinieae could not be reconstructed, but Late Miocene bursts in diversification rates in the Himalayan and southwestern Chinese clades of Aconitum and Delphinium appear to be associated with transitions from short-lived to long-lived life histories.

An experimental test of well-described vegetation patterns across slope aspects using woodland herb transplants and manipulated abiotic drivers.

*The ubiquitous transition of plant communities across slope aspects is a well-described, but rarely tested, ecological dynamic. Aspect position is often used as a proxy for microclimate changes in moisture, light and temperature, but these abiotic drivers are seldom decoupled and very rarely manipulated across slope aspects. *To investigate the mechanisms and demographic stages driving the observed distribution patterns of two woodland herbs in the southeastern USA, seeds and adults were transplanted across north- and south-facing slopes, and moisture and light were experimentally manipulated. *Stage- and species-specific abiotic responses resulted in similar landscape-level patterning for Hexastylis arifolia and Hepatica nobilis, but the underlying abiotic drivers were unique. Adult rather than seed survival best explained the natural distributions across slope aspects, and Hexastylis arifolia was limited by higher temperature, whereas Hepatica nobilis was limited by lower soil moisture. *The stage- and species-specific responses indicated that the use of slope aspect to explain plant distributions not only obfuscates explanatory mechanisms, but probably undermines the portability of results. As abiotic drivers, not topographical proxies, are projected to shift with global climate change, distribution research requires direct abiotic data in association with key demographic stages rather than topographical proxies.

Using microscopic techniques for taxonomic implications of seed and fruits of Delphinium L. (sensu lato) (Ranunculaceae).

In present research, micromorphological characters of the genus Delphinium L. (sensu lato) Ranunculaceae seeds and fruits were studied using microscopic techniques. A total of 37 species were studied using light (LM) and scanning electron microscopy (SEM) to evaluate seeds and fruits micromorphological characteristics for the taxonomic identification and species relationship in the genus. For each taxon, 33 characters were observed and studied. All characters were analyzed using MVSP software and UPGMA method by using dissimilarity index (average taxonomic distance). Considerable differences were observed in trichomes shape of fruit and seed color features. The results revealed that all studied taxa are separated into three clades. The first clade consists of D. pallidiflorum and D. semibarbatum, which have black seed and two types of glandular and crispate trichomes in fruit. The second clade consists of D. speciosum, D. szowitsianum, D. dasystachyson, D. aquilegifolium, D. ursinum, C. trigonelloides, and C. camptocarpa, which had a brown color seeds, two types of glandular and villous trichomes on fruits also have been observed. The remaining species were classified as third clade having various colors including yellow, black, and brown color seeds, the trichome shapes included one of the following forms strigose, villous, trichomes less, glandular, hooked, and crispate trichomes in fruits. The aim of the present study is to use micromorphological characters of seeds and fruits for the taxonomic identification and species delimitation in the genus Delphinium.

Root exudate composition of grass and forb species in natural grasslands.

Plants exude a diverse cocktail of metabolites into the soil as response to exogenous and endogenous factors. So far, root exudates have mainly been studied under artificial conditions due to methodological difficulties. In this study, each five perennial grass and forb species were investigated for polar and semi-polar metabolites in exudates under field conditions. Metabolite collection and untargeted profiling approaches combined with a novel classification method allowed the designation of 182 metabolites. The composition of exuded polar metabolites depended mainly on the local environment, especially soil conditions, whereas the pattern of semi-polar metabolites was primarily affected by the species identity. The profiles of both polar and semi-polar metabolites differed between growth forms, with grass species being generally more similar to each other and more responsive to the abiotic environment than forb species. This study demonstrated the feasibility of investigating exudates under field conditions and to identify the driving factors of exudate composition.

Early reproductive developmental anatomy in Decaisnea (Lardizabalaceae) and its systematic implications.

BACKGROUND AND AIMS: Decaisnea insignis, known as 'dead man's fingers' (Lardizabalaceae), is widely distributed in China and the Himalayan foothill countries. This economically important plant, which is the only species in the genus, has not been the subject of any embryological studies aside from one brief, older paper that lacks micrographs. Data on Decaisnea are also important because its systematic position has been unstable since the genus was established in 1855. Therefore, the objectives of this study were: (a) to use modern microscopy to document early reproductive anatomical development in Decaisnea; and (b) to compare qualitatively these early embryological characters with allied taxa in a systematic context. METHODS: Decaisnea insignis floral buds and inflorescences were regularly collected from Shaanxi Province, China and prepared for light microscopy. The embryological characters studied were qualitatively compared with those of allied taxa via a thorough examination of the existing literature. KEY RESULTS: Early reproductive anatomy in Decaisnea was documented and novel revelations made. It was discovered that the pollen is shed when three-celled (not two-celled, as previously reported), and that endosperm formation is nuclear (not cellular or helobial, as previously reported). These two newly revealed embryological characters are not found in any other members of Lardizabalaceae. Furthermore, neither are persistent antipodal cells, which we confirmed to be present in Decaisnea. CONCLUSIONS: Decaisnea and other Lardizabalaceae characteristically have tetrasporangiate anthers, a secretory tapetum, simultaneous microsporocyte cytokinesis, primarily bitegmic, crassinucellate ovules, and a Polygonum type embryo sac. However, in the family, persistent antipodals, nuclear endosperm, and pollen shed at the three-celled stage are only found in Decaisnea. These embryological data prompted the suggestion that Decaisnea needs elevation above the level of genus.

Structural variation of the complete chloroplast genome and plastid phylogenomics of the genus Asteropyrum (Ranunculaceae).

Two complete chloroplast genome sequences of Asteropyrum, as well as those of 25 other species from Ranunculaceae, were assembled using both Illumina and Sanger sequencing methods to address the structural variation of the cp genome and the controversial systematic position of the genus. Synteny and plastome structure were compared across the family. The cp genomes of the only two subspecies of Asteropyrum were found to be differentiated with marked sequence variation and different inverted repeat-single copy (IR-SC) borders. The plastomes of both subspecies contains 112 genes. However, the IR region of subspecies peltatum carries 27 genes, whereas that of subspecies cavaleriei has only 25 genes. Gene inversions, transpositions, and IR expansion-contraction were very commonly detected in Ranunculaceae. The plastome of Asteropyrum has the longest IR regions in the family, but has no gene inversions or transpositions. Non-coding regions of the cp genome were not ideal markers for inferring the generic relationships of the family, but they may be applied to interpret species relationship within the genus. Plastid phylogenomic analysis using complete cp genome with Bayesian method and partitioned modeling obtained a fully resolved phylogenetic framework for Ranunculaceae. Asteropyrum was detected to be sister to Caltha, and diverged early from subfamily Ranunculoideae.

Genetic variation among wild and cultivated populations of the Chinese medicinal plant Coptis chinensis (Ranunculaceae).

To examine if the cultivation process has reduced the genetic variation of modern cultivars of the traditional Chinese medicinal plant, Coptis chinensis, the levels and distribution of genetic variation was investigated using ISSR markers. A total of 214 C. chinensis individuals from seven wild and three cultivated populations were included in the study. Seven ISSR primers were used and a total of 91 DNA fragments were scored. The levels of genetic diversity in cultivated populations were similar as those in wild populations (mean PPL = 65.2% versus PPL = 52.4%, mean H = 0.159 versus H = 0.153 and mean I = 0.255 versus I = 0.237), suggesting that cultivation did not seriously influence genetic variation of present-day cultivated populations. Neighbour-joining cluster analysis showed that wild populations and cultivated populations were not separated into two groups. The coefficient of genetic differentiation between a cultivar and its wild progenitor was 0.066 (G(st)), which was in good accordance with the result by amova analysis (10.9% of total genetic variation resided on the two groups), indicating that cultivated populations were not genetically differentiated from wild progenitors. For the seven wild populations, a significant genetic differentiation among populations was found using amova analysis (45.9% of total genetic variation resided among populations). A number of causes, including genetic drift and inbreeding in the small and isolated wild populations, the relative limited gene flow between wild populations (N(m) = 0.590), and high gene flow between cultivars and their wild progenitors (N(m) = 7.116), might have led to the observed genetic profiles of C. chinensis.

Identification of green energy ranunculaceous flora of district Chitral, Northern Pakistan using pollen features through scanning electron microscopy.

Pollen micro-morphological features have proven to be helpful for the plant taxonomists in the identification and classification of plants. The utilization of this plantmayhelpfulin the areas of lignocellulosic conversion to biofuels and diversify application toward biomass. The current study was planned with the aim to evaluate the pollen features of complex Ranunculaceous flora of District Chitral, Northern Pakistan using both scanning electron microscopy (SEM) and Light Microscope (LM) for their taxonomic importance. Pollens of 18 Ranunculaceous species belonging to 6 genera were collected from different localities of the research area. SEM and LM were used to examine both qualitative and quantitative micro-morphological features. Sculptring of the sexine include; Scabrate, psilate, echinate, verrucate, perforate gemmate, and reticulate and so forth. Shape of the pollens was sub-spheroidal, spheroidal, prolate, subprolate and oblate and so forth. Type of pollen was ranged from mono to tricolpate and tricolporate. Quantitative characters include length/width of the pollen, colpus, exine thickness, and P/E ratio. Based on these micro-morphological features a taxonomic key was prepared for the fast and correct identification. RESEARCH HIGHLIGHT: Study of the pollen micro-morphological features of Ranunculaceous species by SEM and LM. Analysing both qualitative and quantitative characters of the pollens. Preparation of taxonomic key based on micro-morphological features for the correct and fast identification.

Phylogenetic reassessment of tribe Anemoneae (Ranunculaceae): Non-monophyly of Anemone s.l. revealed by plastid datasets.

Morphological and molecular evidence strongly supported the monophyly of tribe Anemoneae DC.; however, phylogenetic relationships among genera of this tribe have still not been fully resolved. In this study, we sampled 120 specimens representing 82 taxa of tribe Anemoneae. One nuclear ribosomal internal transcribed spacer (nrITS) and six plastid markers (atpB-rbcL, matK, psbA-trnQ, rpoB-trnC, rbcL and rps16) were amplified and sequenced. Both Maximum likelihood and Bayesian inference methods were used to reconstruct phylogenies for this tribe. Individual datasets supported all traditional genera as monophyletic, except Anemone and Clematis that were polyphyletic and paraphyletic, respectively, and revealed that the seven single-gene datasets can be split into two groups, i.e. nrITS + atpB-rbcL and the remaining five plastid markers. The combined nrITS + atpB-rbcL dataset recovered monophyly of subtribes Anemoninae (i.e. Anemone s.l.) and Clematidinae (including Anemoclema), respectively. However, the concatenated plastid dataset showed that one group of subtribes Anemoninae (Hepatica and Anemone spp. from subgenus Anemonidium) close to the clade Clematis s.l. + Anemoclema. Our results strongly supported a close relationship between Anemoclema and Clematis s.l., which included Archiclematis and Naravelia. Non-monophyly of Anemone s.l. using the plastid dataset indicates to revise as two genera, new Anemone s.l. (including Pulsatilla, Barneoudia, Oreithales and Knowltonia), Hepatica (corresponding to Anemone subgenus Anemonidium).

[Preliminary pharmaphylogenetic study on Ranunculaceae].

OBJECTIVE: To deal with the correlation between phylogeny, chemical constituents and pharmaceutical aspects of Ranunculaceae, namely a pharmaphylogenic study of this taxon. METHOD: Based on chemical, pharmaceutical (both ethnopharmacologic and pharmacological) information, linking with different plant systems of Ranunculaceae. RESULT: Chemical constituents of this taxon included several natural groups: benzylisoquinoline alkaloid, ranunculin, triterpenoid glycoside and diterpene alkaloid etc. Ranunculin and magoflorine were found to present simultaneously in some plants of this taxon. CONCLUSION: Combining with therapeutic information, pharmaphylogenic research were in accordance with the phylogenetic system presented by Tamura that Ranunculaceae was proposed to be divided into six sub-families: Helleboroideae, Ranunculoideae, Cimicfugoideae, Isopyroideae, Thalictroideae and Coptidoideae. Results also supported the establishment of Cimicifugoideae.

[Identification study of different Flos Trollii by IR].

OBJECTIVE: To establish a method to identify different Flos Trollii with IR spectra and to provide a new technique for their science estimation and quality identification. METHODS: Their IR spectra were obtained by the method of intuitionistic analysis-selecting point to normalization. RESULTS: Differences on the shape, the position, the height and the relative intensity of absorption peaks were shown in IR spectra of the ten samples. CONCLUSIONS: Different Flos Trollii can be identified using IR spectra obtained by the method of intuitionistic analysis-selecting point to normalization. This new method can be used to study the identification features of the influence factors of Traditional Chinese Medicine quality.

[Pharmaphylogenetic study on isopyroideae (Ranunculaceae)].

OBJECTIVE: To study the pharmaphylogenetic of medicinal plants of Isopyroideae (Ranunculaceae). METHOD: Comprehensively analyze the correlation between phylogeny, chemical constituents and pharmaceutical aspects of Isopyroideae plants, based on chemical, pharmaceutical (both ethnopharmacologic and pharmacological) information, linking with different plant systems of Ranunculaceae. RESULT: Plants from Aquilegia mainly contain flavonoids constituents while the major chemical constituents of Isopyrum are bisbenzylisoquinoline alkaloids. Chemical characteristics also support that this taxon should be separated from Thalictrodeae, and constituted an independent subfamily, namely, Isopyroideae.

Genetical identification of Akebia and Aristolochia species by using Pyrosequencing.

This study was performed to determine if Akebia and Aristolochia species (Akebia quinata Decaisne and Aristolochia manshuriensis Kom.) could be identified by genetic analysis of Pyrosequencing method, which is used to assess genetic variation. The Pyrosequencing results of Akebia and Aristolochia species showed different patterns. The Pyrosequencing analysis of A. quinata Decaisne was very different compared with that of A. manshuriensis Kom. Pyrosequencing analyses might be able to identify the Akebia and Aristolochia species.