The complete annotated plastome sequences of six genera in the tropical woody Polygonaceae.

BACKGROUND: The Polygonaceae is a family well-known for its weeds, and edible plants, Fagopyrum (buckwheat) and Rheum (rhubarb), which are primarily herbaceous and temperate in distribution. Yet, the family also contains a number of lineages that are principally distributed in the tropics and subtropics. Notably, these lineages are woody, unlike their temperate relatives. To date, full-genome sequencing has focused on the temperate and herbaceous taxa. In an effort to increase breadth of genetic knowledge of the Polygonaceae, we here present six fully assembled and annotated chloroplast genomes from six of the tropical, woody genera: Coccoloba rugosa (a narrow and endangered Puerto Rican endemic), Gymnopodium floribundum, Neomillspaughia emarginata, Podopterus mexicanus, Ruprechtia coriacea, and Triplaris cumingiana. RESULTS: These assemblies represent the first publicly-available assembled and annotated plastomes for the genera Podopterus, Gymnopodium, and Neomillspaughia, and the first assembled and annotated plastomes for the species Coccoloba rugosa, Ruprechtia coriacea, and Triplaris cumingiana. We found the assembled chloroplast genomes to be above the median size of Polygonaceae plastomes, but otherwise exhibit features typical of the family. The features of greatest sequence variation are found among the ndh genes and in the small single copy (SSC) region of the plastome. The inverted repeats show high GC content and little sequence variation across genera. When placed in a phylogenetic context, our sequences were resolved within the Eriogonoideae. CONCLUSIONS: These six plastomes from among the tropical woody Polygonaceae appear typical within the family. The plastome assembly of Ruprechtia coriacea presented here calls into question the sequence identity of a previously published plastome assembly of R. albida.

Know your enemy: Application of ATR-FTIR spectroscopy to invasive species control.

Extreme weather and globalisation leave our climate vulnerable to invasion by alien species, which have negative impacts on the economy, biodiversity, and ecosystem services. Rapid and accurate identification is key to the control of invasive alien species. However, visually similar species hinder conservation efforts, for example hybrids within the Japanese Knotweed complex.We applied the novel method of ATR-FTIR spectroscopy combined with chemometrics (mathematics applied to chemical data) to historic herbarium samples, taking 1580 spectra in total. Samples included five species from within the interbreeding Japanese Knotweed complex (including three varieties of Japanese Knotweed), six hybrids and five species from the wider Polygonaceae family. Spectral data from herbarium specimens were analysed with several chemometric techniques: support vector machines (SVM) for differentiation between plant types, supported by ploidy levels; principal component analysis loadings and spectral biomarkers to explore differences between the highly invasive Reynoutria japonica var. japonica and its non-invasive counterpart Reynoutria japonica var. compacta; hierarchical cluster analysis (HCA) to investigate the relationship between plants within the Polygonaceae family, of the Fallopia, Reynoutria, Rumex and Fagopyrum genera.ATR-FTIR spectroscopy coupled with SVM successfully differentiated between plant type, leaf surface and geographical location, even in herbarium samples of varying age. Differences between Reynoutria japonica var. japonica and Reynoutria japonica var. compacta included the presence of two polysaccharides, glucomannan and xyloglucan, at higher concentrations in Reynoutria japonica var. japonica than Reynoutria japonica var. compacta. HCA analysis indicated that potential genetic linkages are sometimes masked by environmental factors; an effect that can either be reduced or encouraged by altering the input parameters. Entering the absorbance values for key wavenumbers, previously highlighted by principal component analysis loadings, favours linkages in the resultant HCA dendrogram corresponding to expected genetic relationships, whilst environmental associations are encouraged using the spectral fingerprint region.The ability to distinguish between closely related interbreeding species and hybrids, based on their spectral signature, raises the possibility of using this approach for determining the origin of Japanese knotweed infestations in legal cases where the clonal nature of plants currently makes this difficult and for the targeted control of species and hybrids. These techniques also provide a new method for supporting biogeographical studies.

Protective effect of Rheum ribes extract against lead-induced hepatotoxicity in male rats

Abstract The effects of Rheum ribes on lead acetate levels and hepatic biochemical factors due to lead acetate toxicity were investigated. Forty male Wistar rats were designated into four groups: Control; lead acetate (receiving in drinking water at 0.6 g/L, daily); hydroalcoholic extract groups (200 and 400 mg/kg doses, gavage, once daily). Treatments were conducted for 10 days. On the 11th day, blood samples were collected to measure lead acetate levels and biochemical factors. Liver tissue samples were examined for histopathological changes. Lead serum levels were increased in lead acetate-treated rats (p<0.001). Lead acetate treatment was associated with a significant increase in liver tissue damage (p<0.001), while R. ribes extract prevented liver tissue damage (p<0.05). The levels of alanine aminotransferase and aspartate aminotransferase were significantly lower in the groups lead acetate + extract (two doses) than in the lead acetate group (p<0.001 and P<0.01, respectively), but alkaline phosphatase level, prothrombin time, partial thromboplastin time and international normalized ratio were not different between the lead acetate + extract groups and the lead acetate group. The results showed the inhibitory role of R. ribes on lead-induced hepato-toxicity. The results make Rhubarb a good candidate to protect against the deleterious effect of chronic lead intoxication after complementary studies

Complete plastome sequencing resolves taxonomic relationships among species of Calligonum L. (Polygonaceae) in China.

BACKGROUND: Calligonum (Polygonaceae) is distributed from southern Europe through northern Africa to central Asia, and is typically found in arid, desert regions. Previous studies have revealed that standard DNA barcodes fail to discriminate Calligonum species. In this study, the complete plastid genomes (plastome) for 32 accessions of 21 Calligonum species is sequenced to not only generate the first complete plastome sequence for the genus Calligonum but to also 1) Assess the ability of the complete plastome sequence to discern species within the group, and 2) screen the plastome sequence for a cost-effective DNA barcode that can be used in future studies to resolve taxonomic relationships within the group. RESULTS: The whole plastomes of Calligonum species possess a typical quadripartite structure. The size of the Calligonum plastome is approximately 161 kilobase pairs (kbp), and encodes 113 genes, including 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. Based on ML phylogenetic tree analyses, the complete plastome has higher species identification (78%) than combinations of standard DNA barcodes (rbcL + matK + nrITS, 56%). Five newly screened gene regions (ndhF, trnS-G, trnC-petN, ndhF-rpl32, rpl32-trnL) had high species resolution, where ndhF and trnS-G were able to distinguish the highest proportion of Calligonum species (56%). CONCLUSIONS: The entire plastid genome was the most effective barcode for the genus Calligonum, although other gene regions showed great potential as taxon-specific barcodes for species identification in Calligonum.

Bridging Inter- and Intraspecific Trait Evolution with a Hierarchical Bayesian Approach.

The evolution of organisms is crucially dependent on the evolution of intraspecific variation. Its interactions with selective agents in the biotic and abiotic environments underlie many processes, such as intraspecific competition, resource partitioning and, eventually, species formation. Nevertheless, comparative models of trait evolution neither allow explicit testing of hypotheses related to the evolution of intraspecific variation nor do they simultaneously estimate rates of trait evolution by accounting for both trait mean and variance. Here, we present a model of phenotypic trait evolution using a hierarchical Bayesian approach that simultaneously incorporates interspecific and intraspecific variation. We assume that species-specific trait means evolve under a simple Brownian motion process, whereas species-specific trait variances are modeled with Brownian or Ornstein-Uhlenbeck processes. After evaluating the power of the method through simulations, we examine whether life-history traits impact evolution of intraspecific variation in the Eriogonoideae (buckwheat family, Polygonaceae). Our model is readily extendible to more complex scenarios of the evolution of inter- and intraspecific variation and presents a step toward more comprehensive comparative models for macroevolutionary studies.

Refugial isolation and range expansions drive the genetic structure of Oxyria sinensis (Polygonaceae) in the Himalaya-Hengduan Mountains.

The formation of the Mekong-Salween Divide and climatic oscillations in Pleistocene were the main drivers for the contemporary diversity and genetic structure of plants in the Himalaya-Hengduan Mountains (HHM). To identify the relative roles of the two historical events in shaping population history of plants in HHM, we investigated the phylogeographic pattern of Oxyria sinensis, a perennial plant endemic to the HHM. Sixteen chloroplast haplotypes were identified and were clustered into three phylogenetic clades. The age of the major clades was estimated to be in the Pleistocene, falling into several Pleistocene glacial stages and postdating the formation of the Mekong-Salween Divide. Range expansions occurred at least twice in the early and middle Pleistocene, but the spatial genetic distribution rarely changed since the Last Glacial Maximum. Our results suggest that temporary mountain glaciers may act as barriers in promoting the lineage divergence in O. sinensis and that subsequential range expansions and secondary contacts might reshape the genetic distribution in geography and blur the boundary of population differentiation created in the earlier glacial stages. This study demonstrates that Pleistocene climatic change and mountain glaciers, rather than the Mekong-Salween Divide, play the primary role in shaping the spatial genetic structure of O. sinensis.

Tertiary montane origin of the Central Asian flora, evidence inferred from cpDNA sequences of Atraphaxis (Polygonaceae).

Atraphaxis has approximately 25 species and a distribution center in Central Asia. It has been previously used to hypothesize an origin from montane forest. We sampled 18 species covering three sections within the genus and sequenced five cpDNA spacers, atpB-rbcL, psbK-psbI, psbA-trnH, rbcL, and trnL-trnF. BEAST was used to reconstruct phylogenetic relationship and time divergences, and S-DIVA and Lagrange were used, based on distribution area and ecotype data, for reconstruction of ancestral areas and events. Our results appear compatible with designation of three taxonomic sections within the genus. The generic stem and crown ages were Eocene, approximately 47 Ma, and Oligocene 27 Ma, respectively. The origin of Atraphaxis is confirmed as montane, with an ancestral area consisting of the Junggar Basin and uplands of the Pamir-Tianshan-Alatau-Altai mountain chains, and ancestral ecotype of montane forest. Two remarkable paleogeographic events, shrinkage of the inland Paratethys Sea at the boundary of the late Oligocene and early Miocene, and the time intervals of cooling and drying of global climate from 24 (22) Ma onward likely facilitated early diversification of Atraphaxis, while rapid uplift of the Tianshan Mountains during the late Miocene may have promoted later diversification.

Species identification of Chinese medicinal plant Fallopia multiflora (Thunb.) Haraldson by suppression subtraction hybridization.

Fallopia multiflora (Thunb.) Haraldson, a traditional Chinese medicinal plant, has been extensively used in preparations of herbal medicine, health products and personal hygiene products. However, the clinical safety and efficiency of F. multiflora (Thunb.) Haraldson is impaired because of the existence of various adulterants. In this study, genomic DNA (gDNA) suppression subtraction hybridization (SSH) was used to authenticate F. multiflora (Thunb.) Haraldson from its adulterants. First, differential gDNA fragments between F. multiflora (Thunb.) Haraldson and its most closely related species F. multiflora var. ciliinervis (Nakai) Yonek. & H. Ohashi by SSH were identified. The differential fragments were then hybridized with arrays constructed from multiple whole genomes of several species (adulterants and/or closely related plants) to screen for the unique gDNA fragments representing F. multiflora (Thunb.) Haraldson. The unique gDNA fragments could be used to design species-specific primers for the identification of F. multiflora (Thunb.) Haraldson. Using SSH, we obtained four differential gDNA fragments, and four pairs of primers were designed. The designed primers could differentiate F. multiflora (Thunb.) Haraldson from its adulterants and/or closely related species via PCR. The results confirmed that the SSH is an efficient method for screening and designing species-specific primers.

DNA barcoding of populations of Fallopia multiflora, an indigenous herb in China.

Fallopia multiflora, locally known as Heshouwu, is one of the most important and widely used Chinese medicinal herbs. However, there is still considerable confusion concerning its different provenances. DNA barcoding is a recent aid to taxonomic identification and uses a short standardized DNA region to discriminate plant species. We assessed the applicability of 4 candidate DNA barcodes (matK, rbcL, psbA-trnH, and ITS2) to identify populations of F. multiflora. To our knowledge, this is the first attempt involving the plant kingdom to apply DNA barcoding at a level lower than species. Four DNA loci (matK, rbcL, psbA-trnH, and ITS2) of 105 samples, including the wild F. multiflora distributed in 17 provinces of China and 4 cultivated F. multiflora lines, were amplified by PCR and sequenced. The 4 loci were evaluated by PCR amplification for sequence quality, extent of genetic divergence, DNA barcoding gap, and the ability to discriminate between populations by BLAST1 and Nearest Distance. We found that psbA-trnH was the best barcode, with significant inter-population variability and best potential for identifying F. multiflora. The combination of loci gave better performance for distinguishing populations than a single locus. We recommend using matK + rbcL + psbA-trnH + ITS2 or psbA-trnH alone for this species. This research demonstrates the utility of DNA barcoding for geoherbalism identifications.

Molecular phylogeny of Koenigia L. (Polygonaceae: Persicarieae): implications for classification, character evolution and biogeography.

To examine the phylogenetic relationships of Koenigia sensu lato (Polygonaceae), 43 samples representing all species of Koenigia and closely related taxa (e.g., Aconogonon, Bistorta, and Persicaria) were sequenced for nuclear ITS and four plastid regions (trnL-F, atpB-rbcL, rbcL, and rpl32-trnL((UAG))). Phylogenetic analyses indicate that Koenigia recognized by Hedberg is paraphyletic and that the basal species K. delicatula should be reassigned to a separate new genus. Based on these findings, we further propose that the genus Koenigia sensu lato be circumscribed to include five species. Ancestral state reconstruction showed that the pollen apertures likely evolved in parallel in the Aconogonon-Koenigia-Bistorta clade and Persicaria clade and that tricolpate pollen is most likely to be the ancestral state. Quincuncial aestivation likely evolved during the early evolution of Koenigia and its close relatives. Our findings suggest that the uplift of the Himalayas has played an important role in promoting species diversification of Koenigia. Koenigia islandica expanded its range during Pleistocene glacial cycles by tracking changes in newly available habitats.

Age estimates for the buckwheat family Polygonaceae based on sequence data calibrated by fossils and with a focus on the amphi-Pacific Muehlenbeckia.

The buckwheat family Polygonaceae is a diverse group of plants and is a good model for investigating biogeography, breeding systems, coevolution with symbionts such as ants and fungi, functional trait evolution, hybridization, invasiveness, morphological plasticity, pollen morphology and wood anatomy. The main goal of this study was to obtain age estimates for Polygonaceae by calibrating a Bayesian phylogenetic analysis, using a relaxed molecular clock with fossil data. Based on the age estimates, we also develop hypotheses about the historical biogeography of the Southern Hemisphere group Muehlenbeckia. We are interested in addressing whether vicariance or dispersal could account for the diversification of Muehlenbeckia, which has a "Gondwanan" distribution. Eighty-one species of Polygonaceae were analysed with MrBayes to infer species relationships. One nuclear (nrITS) and three chloroplast markers (the trnL-trnF spacer region, matK and ndhF genes) were used. The molecular data were also analysed with Beast to estimate divergence times. Seven calibration points including fossil pollen and a leaf fossil of Muehlenbeckia were used to infer node ages. Results of the Beast analyses indicate an age of 110.9 (exponential/lognormal priors)/118.7 (uniform priors) million years (Myr) with an uncertainty interval of (90.7-125.0) Myr for the stem age of Polygonaceae. This age is older than previously thought (Maastrichtian, approximately 65.5-70.6 Myr). The estimated divergence time for Muehlenbeckia is 41.0/41.6 (39.6-47.8) Myr and its crown clade is 20.5/22.3 (14.2-33.5) Myr old. Because the breakup of Gondwana occurred from 95-30 Myr ago, diversification of Muehlenbeckia is best explained by oceanic long-distance and maybe stepping-stone dispersal rather than vicariance. This study is the first to give age estimates for clades of Polygonaceae and functions as a jumping-off point for future studies on the historical biogeography of the family.

[Study on origin and species of Heshouwu].

Recently the safety of Heshouwu become a focus, but the reasons of its hepotoxicity are confused. On the basis of literature research, some findings on species and usage custom maybe supply some clues to explain the reasons of its hepotoxicity. Heshouwu had red Heshouwu (male) and white Heshouwu (female) in ancient literature, and traditional usage was use of the male and female together. The Latin name of red Heshouwu is Fallopia multiflora, and that of the white one is F. multiflora var. multiflora.

[Molecular authentication of Fallopia multiflora by PCR-RFLP based on the trnL-trnF analysis].

OBJECTIVE: To establish a method for the molecular authentication of Fallopia multiflora. METHODS: The trnL-trnF regions of Fallopia multiflora and its closely related species and/or adulterants were sequenced and analyzed. RESULTS: It was found that the trnL-trnF sequence divergences between Fallopia multiflora and its closely related species and/or adulterants were 2.1%-22%. While the intra-species trnL-trnF divergences of Fallopia multiflora were 0%-1.5%. Based on the trnL-trnF regional variations, an endonuclease Xba I (T CTAGA) restriction site specific to Fallopia multiflora was detected. The Fallopia multiflora trnL-F polymerase chain reaction product could be cleaved by Xba I into two pieces, 804-819 bp and 256 bp each, whereas the restriction endonuclease could not digest the trnL-trnF polymerase chain reaction product of its closely related species or adulterants. The restriction patterns analyzed for restriction enzyme Xba I were found to be identical in all Fallopia multiflora individuals from different geographical regions in China. CONCLUSION: The assay based on polymerase chain reaction amplification of the trnL-trnF fragment of chloroplast DNA and subsequent restriction fragment length polymorphism can be used as a general test to identify Fallopia multiflora.

Chemical and genetic diversity among some wild stands of Calligonum polygonoides (Polygonaceae) from the Thar Desert of Rajasthan.

The arid Western Rajasthan, where the Thar Desert of India is immersed, is mostly covered by sand dunes, a common landscape. The region has confronted with fragilities of natural resources, low, erratic and ill-distributed rainfall, and is covered up with many perennial hardy shrubs. Calligonumpolygonoides, the most common perennial shrub, is widely present in some localities of this Thar Desert. In this study, we evaluated the diversity present among 54 wild Calligonum polygonoides plants, sampled from eight different locations within the Thar Desert. Our methods included chemical/nutritional characteristics and random amplified polymorphic DNA (RAPD). Both chemical and molecular methods produced wider range of diversity, however, RAPD detected comparatively more diversity. A total of 163 band positions were produced by ten RAPD primers, of which 147 were found polymorphic with 90.18% polymorphism. RAPD-based Jaccard's similarity coefficients ranged from 0.43-0.89. The analysis of various chemical and mineral constituents revealed that phog is an excellent source of calcium, potassium and phosphorous while relatively poor in zinc. Among minerals, average potassium content was found maximum (2 430mg/100g) with 0.14 CV. Zinc was observed comparably less in quantity while highest variable with CV 0.73. The chemical-based Manhattan dissimilarity coefficient values ranged from 0.01-0.22 with an average of 0.12. The comparison of the clusters obtained based on the chemical and mineral parameters with those of the RAPD data showed that the groups formed in both cases showed different patterns of relationships among the samples. Broader range of diversity might be due to the out breeding behavior of C. polygonoides and indicates the good adaptability of the plants in the region studied. However, low diversity observed in the Bikaner province is alarming and suggests that anthropogenic activities leading to heavy population disturbances can affect the genetic composition of the species in a considerable way.

Chemical and genetic diversity among some wild stands of Calligonum polygonoides (Polygonaceae) from the Thar Desert of Rajasthan

The arid Western Rajasthan, where the Thar Desert of India is immersed, is mostly covered by sand dunes, a common landscape. The region has confronted with fragilities of natural resources, low, erratic and ill-distributed rainfall, and is covered up with many perennial hardy shrubs. Calligonum polygonoides, the most common perennial shrub, is widely present in some localities of this Thar Desert. In this study, we evaluated the diversity present among 54 wild Calligonum polygonoides plants, sampled from eight different locations within the Thar Desert. Our methods included chemical/nutritional characteristics and random amplified polymorphic DNA (RAPD). Both chemical and molecular methods produced wider range of diversity, however, RAPD detected comparatively more diversity. A total of 163 band positions were produced by ten RAPD primers, of which 147 were found polymorphic with 90.18% polymorphism. RAPD-based Jaccard’s similarity coefficients ranged from 0.43-0.89. The analysis of various chemical and mineral constituents revealed that phog is an excellent source of calcium, potassium and phosphorous while relatively poor in zinc. Among minerals, average potassium content was found maximum (2 430mg/100g) with 0.14 CV. Zinc was observed comparably less in quantity while highest variable with CV 0.73. The chemical-based Manhattan dissimilarity coefficient values ranged from 0.01-0.22 with an average of 0.12. The comparison of the clusters obtained based on the chemical and mineral parameters with those of the RAPD data showed that the groups formed in both cases showed different patterns of relationships among the samples. Broader range of diversity might be due to the out breeding behavior of C. polygonoides and indicates the good adaptability of the plants in the region studied. However, low diversity observed in the Bikaner province is alarming and suggests that anthropogenic activities leading to heavy population disturbances can affect the genetic composition of the species in a considerable way.

El árido Rajastán occidental, en donde está inmerso el desierto de Thar en la India, está cubierto principalmente por dunas de arena, un paisaje común. La región ha enfrentado la fragilidad de los recursos naturales, las lluvias escasas, irregulares y mala distribución, y está cubierta con muchos arbustos resistentes perennes. Calligonum polygonoides, el arbusto perenne más común, se encuentra ampliamente en algunas localidades del desierto de Thar. En este estudio, se evaluó la diversidad presente entre 54 plantas silvestres de Calligonum polygonoides, de ocho localidades diferentes del desierto de Thar. Nuestros métodos incluyen características químicas/nutricionales y ADN polimórfico amplificado (RAPD) al azar. Ambos métodos químicos y moleculares producen un amplio rango de la diversidad, sin embargo, RAPD detectó comparativamente mayor diversidad. Un total de 163 posiciones de la banda fueron producidos por diez cebadores RAPD, de los cuales 147 se encontraron polimórficos con un 90.18% de polimorfismo. El coeficiente de RAPD basado en la similitud de Jaccard varió desde 0.43 hasta 0.89. El análisis de varios constituyentes químicos y minerales reveló que Calligonum polygonoides es una excelente fuente de calcio, potasio y fósforo mientras que es relativamente pobre en zinc. Entre los minerales, el contenido de potasio promedio se encontró como máximo (2 430mg/100g), con 0.14 CV. El zinc se observó comparativamente menor en cantidad, pero presentó la mayor variabilidad con CV 0.73. El valor del coefficente de disimilitud de Manhattan varió en un rango de 0.01 hasta 0.22 con un promedio de 0.12. La comparación de los grupos obtenidos según los parámetros químicos y minerales con las de los datos de RAPD mostró que los grupos formados en ambos casos mostraron patrones diferentes de relaciones entre las muestras. Una gama más amplia de la diversidad podría ser debido al comportamiento reproductivo C polygonoides e indica la buena adaptabilidad ...

Taxa distribution and RAPD markers indicate different origin and regional differentiation of hybrids in the invasive Fallopia complex in central-western Europe.

Interspecific hybridization can be a driving force for evolutionary processes during plant invasions, by increasing genetic variation and creating novel gene combinations, thereby promoting genetic differentiation among populations of invasive species in the introduced range. We examined regional genetic structure in the invasive Fallopia complex, consisting of F. japonica var. japonica, F. sachalinensis and their hybrid F. x bohemica, in seven regions in Germany and Switzerland using RAPD analysis and flow cytometry. All individuals identified as F. japonica var. japonica had the same RAPD phenotype, while F. sachalinensis (11 RAPD phenotypes for 11 sampled individuals) and F. x bohemica (24 RAPD phenotypes for 32 sampled individuals) showed high genotypic diversity. Bayesian cluster analysis revealed three distinct genetic clusters. The majority of F. x bohemica individuals were assigned to a unique genetic cluster that differed from those of the parental species, while the other F. x bohemica individuals had different degrees of admixture to the three genetic clusters. At the regional scale, the occurrence of male-fertile F. sachalinensis coincided with the distribution of F. x bohemica plants showing a high percentage of assignment to both parental species, suggesting that they originated from hybridization between the parental species. In contrast, in regions where male-fertile F. sachalinensis were absent, F. x bohemica belonged to the non-admixed genetic group, indicating multiple introductions of hybrids or sexual reproduction among hybrids. We also found regional differentiation in the gene pool of F. x bohemica, with individuals within the same region more similar to each other than to individuals from different regions.

[Analysis and authentication of Fallopia multiflora and its adulterants by matK sequence].

OBJECTIVE: To identify Fallopia multiflora from its adulterants by comparing their matK sequences. METHODS: Genomic DNA of different materials was extracted using modified cetytrimethyl ammonium bromide (CTAB) method. The double-strand matK genes were amplified using PCR method and then sequenced. The data were analyzed in Clustral W and MEGA 4.0 software package. RESULTS: Besides F. multiflora var. ciliinerve, the matK sequences of other adulterants show distinct differences with F. multiflora, whether for nucleotides substitutions or genetic distances; and the specific identifying sites for distinguishing F. multiflora and other Fallopia adulterants were found through further comparative analysis. Moreover, the 3 inspected materials were successfully authenticated by comparing the matK sequences. CONCLUSION: matK sequences can be used for the molecular identification between F. multiflora and its adulterant species.

Authentication of the family Polygonaceae in Chinese pharmacopoeia by DNA barcoding technique.

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants belonging to the family Polygonaceae in Chinese pharmacopoeia possess important medicinal efficacy in traditional Chinese medicines. AIM OF THE STUDY: DNA barcodes are first used to discriminate the Polygonaceae in Chinese pharmacopoeia and their adulterants. MATERIALS AND METHODS: DNA samples, extracted from thirty-eight specimens belonging to eighteen species in Polygonaceae, were used as templates. Eight candidate barcodes were amplified by polymerase chain reaction. Sequence analysis was accomplished by CodonCode Aligner V 2.06 and DNAman V 6. Species identification was performed using MEGA V 4.0. RESULTS: The amplification efficiency of six candidate DNA barcodes (rbcL, trnH-psbA, ndhJ, rpoB, rpoC1, accD) was 100%, while the efficiency of YCF5 and nrITS was 56% and 44%, respectively. The interspecific divergence was highest for the trnH-psbA (20.05%), followed by the nrITS (14.01%) across all species pairs, while intraspecific variation both within populations and between populations was absent (0.0%). The trnH-psbA can not only distinguish ten species of Polygonaceae in Chinese pharmacopoeia, but also recognize eight other species of Polygonaceae including their adulterants. CONCLUSION: Our findings show that DNA barcoding is an efficient tool for identification of Polygonaceae in Chinese pharmacopoeia and their adulterants.

Genetic variation and identification of cultivated Fallopia multiflora and its wild relatives by using chloroplast matK and 18S rRNA gene sequences.

FALLOPIA MULTIFLORA (Thunb.) Harald . has been widely and discriminatingly used in China for the study and treatment of anemia, swirl, deobstruent, pyrosis, insomnia, amnesia, atheroma and also for regulating immune functions. However, there is still confusion about the herbal drug's botanical origins and the phylogenetic relationship between the cultivars and the wild relatives. In order to develop an efficient method for identification, a molecular analysis was performed based on 18 S rRNA gene and partial MATK gene sequences. The 18 S rRNA gene sequences of F. MULTIFLORA were 1809 bp in length and were highly conserved, indicating that the cultivars and the wild F. MULTIFLORA have the same botanical origin. Based on our 18 S rRNA gene sequences analysis, F. MULTIFLORA could be easily distinguished at the DNA level from adulterants and some herbs with similar components. The MATK gene partial sequences were found to span 1271 bp. The phylogenetic relation of F. MULTIFLORA based on the MATK gene showed that all samples in this paper were divided into four clades. The sequences of the partial MATK gene had many permutations, which were related to the geographical distributions of the samples. MATK gene sequences provided valuable information for the identification of F. MULTIFLORA. New taxonomic information could be obtained to authenticate the botanical origin of the F. MULTIFLORA, the species and the medicines made of it.

Simultaneous qualitative and quantitative analysis of commercial bistorta rhizome and its differentiation from closely related herbs using TLC and HPLC-DAD fingerprinting.

A qualitative and quantitative analysis method was established to improve quality assessment standards for Rhizoma Polygoni Bistortae (Polygonum bistorta L.) and differentiate commercial bistorta rhizome from closely related herbs by TLC and HPLC-DAD fingerprinting. Three compounds including phenolic acid and flavane were identified by comparison with standard compounds and quantified simultaneously by HPLC-DAD simultaneously. A comprehensive validation of the method that included sensitivity, linearity, repeatability and recovery was conducted. Paris polyphylla SM., a herb often mixed with Polygonum bistorta L. in China due to their same popular name "Caoheche" in history, was successfully distinguished by thin-layer chromatography (TLC) fingerprinting of the petroleum-soluble fraction. Polygonum paleaceum WALL., another herb often mixed with Polygonum bistorta L. due to their similar external appearances, was distinguished by HPLC fingerprinting.