Non-coding Regions Of Chloroplast DNA Research Articles

Universal PCR primers for identifying species of the genus Sanguisorba and other members of the family Rosaceae

Seven primers for the amplification of non-coding regions of chloroplast DNA via the polymerase chain reaction (PCR) have been designed. In order to find out whether these primers were universal, we used them in an attempt to amplify DNA fragments from various plant species of Rosaceae. Primers worked for species of the genus Sanguisorba and some species of the family Rosaceae. The fact that they amplify chloroplast DNA non-coding regions over a wide taxonomic range means that these primers may be used to study the population biology and evolution of plants.

New Approach to the Systematics of the Section Psammiris (Iris, Iridaceae): What Does Chloroplast DNA Sequence Tell Us?

Iris sect. Psammiris comprises rhizomatous perennials distributed in the north temperate zone of Eurasia. The systematics of the section are currently based on morphology, and the phylogenetic relationships within it still remain unclear. In the framework of Iris systematics, we conducted molecular and morphological analyses of the currently recognized I. sect. Psammiris species to elucidate the taxonomic composition and relationships within the section. The phylogenetic reconstructions based on sequence variation of four noncoding chloroplast DNA regions support the monophyly of I. sect. Psammiris, which includes I. tigridia, while I. potaninii var. ionantha belongs to I. sect. Pseudoregelia. The proposed novel classification of I. sect. Psammiris recognizes three series: an autonymic series with I. humilis, I. bloudowii, and I. vorobievii and two unispecific series (I. ser. Potaninia with I. potaninii and I. ser. Tigridiae with I. tigridia). In addition, the taxonomic statuses of I. arenaria, I. ivanovae, I. kamelinii, I. mandshurica, I. pineticola, I. psammocola, and I. schmakovii are clarified herein. We provide a revised taxonomic treatment for I. sect. Psammiris, including notes on the types; updated information on species synonymy, distributions, habitats, and chromosome numbers; and a new identification key to the species. Three lectotypes are designated here.

The role of seaweed extract on improvement drought tolerance of wheat revealed by osmoprotectants and DNA (cpDNA) markers

Drought stress is one of the most severe abiotic stresses affecting adversely plant growth, crop production, and various metabolic processes. Using seaweed extract in mitigating water stress adverse effects is highly important for plant production. The present study discussed the physiological role of seaweed extract (Sargassum denticulatum) in improving wheat tolerance to water stress.Water stress (40% of field capacity) caused significant decreases in wheat plant growth parameters (shoot height, fresh, and dry weights of the shoot) as well as with significant decreases in chlorophyll content and starch. Total soluble sugars, free amino acids, proline, and phenolic compounds contents increased in stressed wheat plants irrigated every three weeks compared with control plants. The foliar application of seaweed extract 2% enhanced all growth and yield parameters and more accumulation of the organic solutes in leaves of water-stressed plants. These increases correlated with significant increases in total phenolic contents as compared with control plants. The trnL intron and psbA-trnH intergenic regions of cpDNA were amplified from extracted total genomic DNA. The results indicated that the variation among psbA-trnH intergenic region was more than trnL intron region to distinct the variation of wheat treatments as responsible to water deficit.Foliar spray of seaweeds extract was effective in improving wheat performance by enhancing compatible osmolytes, antioxidant compounds and enhancing variation among non-coding chloroplast DNA (cpDNA) regions trnL intron and psbA-tnH as a response to water deficit.

Erianthus germplasm collection in Thailand: genetic structure and phylogenetic aspects of tetraploid and hexaploid accessions

BackgroundThe genus Erianthus, which belongs to the “Saccharum complex”, includes C4 warm-season grasses. Erianthus species are widely distributed throughout Southeast Asia, East Asia and South Asia. Erianthus arundinaceus (Retz.) Jeswiet is highly adaptable to the environment, has a high percentage of dry matter, and is highly productive. Recently, this species has attracted attention as a novel bioenergy crop and as a breeding material for sugarcane improvement. Such interest in E. arundinaceus has accelerated the collection and conservation of its genetic resources, mainly in Asian countries, and also evaluation of morphological, agricultural, and cytogenetic features in germplasm collections. In Thailand, genetic resources of E. arundinaceus have been collected over the past 20 years and their phenotypic traits have been evaluated. However, the genetic differences and relatedness of the germplasms are not fully understood.ResultsA set of 41 primer pairs for nuclear simple sequence repeats (SSRs) developed from E. arundinaceus were used to assess the genetic diversity of 121 Erianthus germplasms collected in Thailand; of these primer pairs, 28 detected a total of 316 alleles. A Bayesian clustering approach with these alleles classified the accessions into four main groups, generally corresponding to the previous classification based on phenotypic analysis. The results of principal coordinate analysis and phylogenetic analysis of the 121 accessions on the basis of the SSR markers showed the same trend as Bayesian clustering, whereas sequence variations of three non-coding regions of chloroplast DNA revealed eight haplotypes among the accessions. The analysis of genetic structure and phylogenetic relationships, however, found some accessions whose classification contradicted the results of previous phenotypic classification.ConclusionsThe molecular approach used in this study characterized the genetic diversity and relatedness of Erianthus germplasms collected across Thailand. This knowledge would allow efficient maintenance and conservation of the genetic resources of this grass and would help to use Erianthus species as breeding materials for development of novel bioenergy crops and sugarcane improvement.

How co-distribution of two related azaleas (Rhododendron) developed in the Japanese archipelago: insights from evolutionary and demographic analyses

Plants growing in similar climates with extensively overlapping distributions may exhibit similar phylogeographic patterns, due to similarities in range shifts during past climatic oscillations. We tested and exploited this expectation in a comparison of the evolutionary and demographic histories of two related and co-distributed species endemic to Japan—Rhododendron pentaphyllum and R. quinquefolium. Genetic variation in sequences of noncoding chloroplast DNA regions and hundreds of nuclear single nucleotide polymorphisms were used to investigate 191 individuals from 13 populations of R. pentaphyllum and 142 individuals from 10 populations of R. quinquefolium. We found that the two species were not each other’s closest relative, and we found different times to the most recent common ancestor of extant populations based on both chloroplast DNA and nuclear SNPs. Rhododendron pentaphyllum exhibited higher genetic variation between populations than R. quinquefolium, while the two species had similar patterns of genetic divergence between regions with slightly different divergence times. These lines of evidence suggest that the similar ranges of the two species are the result of repeated range shifts since speciation during climatic oscillations.

SPInDel Analysis of the Non-Coding Regions of cpDNA as a More Useful Tool for the Identification of Rye (Poaceae: Secale) Species.

Secale is a small but very diverse genus from the tribe Triticeae (family Poaceae), which includes annual, perennial, self-pollinating and open-pollinating, cultivated, weedy and wild species of various phenotypes. Despite its high economic importance, classification of this genus, comprising 3–8 species, is inconsistent. This has resulted in significantly reduced progress in the breeding of rye which could be enriched with functional traits derived from wild rye species. Our previous research has suggested the utility of non-coding sequences of chloroplast and mitochondrial DNA in studies on closely related species of the genus Secale. Here we applied the SPInDel (Species Identification by Insertions/Deletions) approach, which targets hypervariable genomic regions containing multiple insertions/deletions (indels) and exhibiting extensive length variability. We analysed a total of 140 and 210 non-coding sequences from cpDNA and mtDNA, respectively. The resulting data highlight regions which may represent useful molecular markers with respect to closely related species of the genus Secale, however, we found the chloroplast genome to be more informative. These molecular markers include non-coding regions of chloroplast DNA: atpB-rbcL and trnT-trnL and non-coding regions of mitochondrial DNA: nad1B-nad1C and rrn5/rrn18. Our results demonstrate the utility of the SPInDel concept for the characterisation of Secale species.

A littoral grass growing inland: genetic diversity of Puccinellia fasciculata around mud volcanoes in Italy

Puccinellia fasciculata is a littoral miohalophyte with a Mediterranean-Atlantic distribution, uncommonly found in inland sites. The main Italian inland population is located in the Salse di Nirano Reserve, an area internationally renowned for the phenomenon of mud volcanoes. In this study, molecular markers are used to characterize this population and its possible relationships with the nearest littoral conspecific population. A survey of ISSR markers revealed low levels of genetic diversity (He = 0.081 in the inland population, 0.105 in the littoral one) and a weak genetic differentiation (Gst = 0.24 within the inland population, 0.28 among this and the littoral one). A sequence analysis of three non-coding regions of chloroplast DNA found no genetic differentiation both within the inland population and between the two populations, and revealed a common origin for the two dating back to the middle Holocene. The apparent incongruence between the results from the two approaches may be explained by differences between ISSR and non-coding cpDNA markers in capturing signatures related to gene flow; their integrated information implies a mixed reproductive strategy and a common evolutionary history for the two examined populations. Effective conservation strategies are recommended for the inland population of P. fasciculata and its habitat.

Chloroplast and Nuclear Ribosomal DNA Variation in Two Geographically and Ecologically Overlapping Hedysarum Species

We studied the nucleotide variability in two morphologically similar and taxonomically close endemic legume species, Hedysarum austrosibiricum and H. consanguineum. To estimate the level of genetic divergence between the species, four DNA loci were sequenced and analysed: ITS region of nuclear ribosomal DNA and three non-coding regions of chloroplast DNA, viz. trnL-trnF, trnHGUG-psbA, and rpl32-trnLUAG. The chloroplast marker rpl32-trnL was studied in Hedysarum for the first time. Probability of introgressive hybridization between H. austrosibiricum and H. consanguineum was assessed in 17 accessions representing two sympatric and eight allopatric natural populations. The results show that an accession of H. consanguineum contained two trnL-F chlorotypes indicating its heteroplasmic nature. Based on our results, persistence of ancestral polymorphism in Hedysarum is partly present in this species due to the genetic drift and incomplete sorting of the gene lineages.

Dispersal as a result of asymmetrical hybridization between two closely related oak species in China

Hybridization has played an important role in plant evolution. Less attention has been paid, however, to its role in dispersal. In this study, historical divergence and hybridization were investigated in two closely related Chinese oaks, Quercus mongolica and Q. liaotungensis, to estimate the role that hybridization played in their dispersal. We genotyped 27 Q. mongolica and Q. liaotungensis populations throughout the distributional range of the two oak species, using 14 single-copy nuclear genes and four noncoding chloroplast DNA regions. Bayesian cluster and population tree analyses indicated that there were three groups over all oak populations, namely, Q. mongolica, northwest–northern China (NW-NC) Q. liaotungensis, and northeastern China (NEC) Q. liaotungensis. Approximate Bayesian computation simulation supported an asymmetrical hybridization origin of NEC Q. liaotungensis, after a previous divergence between NW-NC Q. liaotungensis and Q. mongolica. IMa3 analyses suggested that Q. liaotungensis and Q. mongolica diverged in the NW-NC and NEC regions, respectively, and that NEC Q. liaotungensis arose from Q. mongolica, not from NW-NC Q. liaotungensis, and was greatly introgressed by NW-NC Q. liaotungensis. Oak populations in NW-NC and NEC regions held different chloroplast DNA haplotypes, and Q. liaotungensis in NEC shared most haplotypes with Q. mongolica populations, but none with NW-NC Q. liaotungensis populations, suggesting the maternal origin of NEC Q. liaotungensis from Q. mongolica. This study found clear signals of isolation divergence of Q. liaotungensis in NW-NC and Q. mongolica in NEC, and the results suggest that asymmetrical hybridization and introgression from Q. liaotungensis to Q. mongolica, mostly likely via pollen flow, facilitated Q. liaotungensis dispersal to NEC.

Applied phylogeography of Cyclopia intermedia (Fabaceae) highlights the need for 'duty of care' when cultivating honeybush.

BackgroundThe current cultivation and plant breeding of Honeybush tea (produced from members of CyclopiaVent.) do not consider the genetic diversity nor structuring of wild populations. Thus, wild populations may be at risk of genetic contamination if cultivated plants are grown in the same landscape. Here, we investigate the spatial distribution of genetic diversity within Cyclopia intermedia E. Mey.—this species is widespread and endemic in the Cape Floristic Region (CFR) and used in the production of Honeybush tea.MethodsWe applied High Resolution Melt analysis (HRM), with confirmation Sanger sequencing, to screen two non-coding chloroplast DNA regions (two fragments from the atpI-aptH intergenic spacer and one from the ndhA intron) in wild C. intermedia populations. A total of 156 individuals from 17 populations were analyzed for phylogeographic structuring. Statistical tests included analyses of molecular variance and isolation-by-distance, while relationships among haplotypes were ascertained using a statistical parsimony network.ResultsPopulations were found to exhibit high levels of genetic structuring, with 62.8% of genetic variation partitioned within mountain ranges. An additional 9% of genetic variation was located amongst populations within mountains, suggesting limited seed exchange among neighboring populations. Despite this phylogeographic structuring, no isolation-by-distance was detected (p > 0.05) as nucleotide variation among haplotypes did not increase linearly with geographic distance; this is not surprising given that the configuration of mountain ranges dictates available habitats and, we assume, seed dispersal kernels.ConclusionsOur findings support concerns that the unmonitored redistribution of Cyclopia genetic material may pose a threat to the genetic diversity of wild populations, and ultimately the genetic resources within the species. We argue that ‘duty of care’ principles be used when cultivating Honeybush and that seed should not be translocated outside of the mountain range of origin. Secondarily, given the genetic uniqueness of wild populations, cultivated populations should occur at distance from wild populations that is sufficient to prevent unintended gene flow; however, further research is needed to assess gene flow within mountain ranges.

Interspecific Hybridization and Introgression Influence Biodiversity—Based on Genetic Diversity of Central European Viola epipsila-V. palustris Complex

The Viola epipsila-V. palustris complex is a highly taxonomically complicated group of species in its entire circumboreal range of distribution. Habitat loss, forest flooding, and hybridization could lead to the extinction of V. epipsila. A hybrid index and principal component analysis (PCA) were used to select qualitative and quantitative morphological features to distinguish parent species and hybrids, inter simple sequence repeat (ISSR) markers to determine the genetic diversity of the populations, flow cytometry to estimate the genome size (GS), and non-coding chloroplast DNA (cpDNA) regions to indicate the directions of crosses. All taxa are very morphologically variable, and their features can change within a season. The most stable feature is the distance of the bracts on the pedicel from the rhizome. The genetic diversity of all taxa populations is low and highly influenced by selfing and vegetative propagation. The population structure is differentiated: populations of V. epipsila or V. palustris, mixed populations with both parent species, F1 hybrids and populations with introgressive forms occur in different regions. The interspecific GS variation corresponds to the ploidy level (4x = 2.52 pg, 8x = 4.26 pg, 6x = 3.42 pg). Viola epipsila is the mother plant of the hybrids. Research has shown the risk of V. epipsila extinction in Central Europe and the importance of local populations in studying the role of hybridization in reducing/maintaining/increasing biodiversity.

Genetic diversity and population structure of the endangered orchid Pelatantheria scolopendrifolia (Orchidaceae) in Korea.

Due to substantial population decline, the Korean orchid P. scolopendrifolia is considered endangered and highly threatened. Like many endangered species, it is vulnerable to biological and anthropogenic threats that can lead to the loss of genetic diversity and, ultimately, extinction. Therefore, the assessment of genetic diversity and population genetic structure is imperative for conservation. In this study, we newly developed 15 polymorphic microsatellite markers. Analyses of genetic diversity and population genetic structure that included 182 samples from 11 populations were conducted using microsatellite markers and four noncoding regions of chloroplast DNA. Our study revealed a relatively low level of genetic diversity (Ho = 0.529, He = 0.356), albeit harboring with private alleles based on microsatellite genotyping data, and high haplotype diversities based on chloroplast DNA sequences data. The results of STRUCTURE and PCoA based on microsatellite genotyping data showed population differentiations. An AMOVA based on chloroplast DNA sequence data further corroborated these conclusions, indicating about 70% of variations found among populations. Low genetic diversity and divergence among the population might have been caused by factors, such as asexual reproduction, demographic events (bottleneck and population expansion), geographic isolation, and low gene flow. The development and implication of conservation strategies and management of P. scolopendrifolia are proposed based on these results.

Genetic Diversity and Demographic History of an Upper Hill Dipterocarp (Shorea platyclados): Implications for Conservation.

Southeast Asian rainforests at upper hill elevations are increasingly vulnerable to degradation because most lowland forest areas have been converted to different land uses. As such, understanding the genetics of upper hill species is becoming more crucial for their future management and conservation. Shorea platyclados is an important, widespread upper hill dipterocarp in Malaysia. To elucidate the genetic structure of S. platyclados and ultimately provide guidelines for a conservation strategy for this species, we carried out a comprehensive study of the genetic diversity and demographic history of S. platyclados. Twenty-seven populations of S. platyclados across its range in Malaysia were genotyped at 15 polymorphic microsatellite loci and sequenced at seven noncoding chloroplast DNA (cpDNA) regions. A total of 303 alleles were derived from the microsatellite loci, and 29 haplotypes were identified based on 2892 bp of concatenated cpDNA sequences. The populations showed moderately high genetic diversity (mean HE = 0.680 for microsatellite gene diversity and HT = 0.650 for total haplotype diversity) and low genetic differentiation (FST = 0.060). Bayesian clustering divided the studied populations into two groups corresponding to western and eastern Malaysia. Bottleneck analysis did not detect any recent bottleneck events. Extended Bayesian skyline analyses showed a model of constant size for the past population history of this species. Based on our findings, priority areas for in situ and ex situ conservation and a minimum population size are recommended for the sustainable utilization of S. platyclados.

Multiple origins and the population genetic structure of Rubus takesimensis (Rosaceae) on Ulleung Island: Implications for the genetic consequences of anagenetic speciation.

To determine the origin and genetic consequences of anagenesis in Rubus takesimensis on Ulleung Island, Korea, we compared the genetic diversity and population structure of R. takesimensis with those of its continental progenitor R. crataegifolius. We broadly sampled a total of 315 accessions in 35 populations and sequenced five noncoding regions of chloroplast DNA. Rubus takesimensis emerged as nonmonophyletic and several geographically diverse continental populations were likely responsible for the origin of R. takesimensis; the majority of R. takesimensis accessions were sisters to the clade containing accessions of R. crataegifolius, primarily from the Korean peninsula, while rare accessions from three populations shared common ancestors with the ones from the southern part of the Korean peninsula, Jeju Island, and Japan. A few accessions from the Chusan population originated independently from the Korean peninsula. Of 129 haplotypes, 81 and 48 were found exclusively in R. crataegifolius and R. takesimensis, respectively. We found unusually high genetic diversity in two regions on Ulleung Island and no geographic population structure. For R. crataegifolius, two major haplotype groups were found; one for the northern mainland Korean peninsula, and the other for the southern Korean peninsula and the Japanese archipelago. Compared with populations of R. crataegifolius sampled from Japan, much higher haplotype diversity was found in populations from the Korean peninsula. The patterns of genetic consequences in R. takesimensis need to be verified for other endemic species based on chloroplast DNA and independent nuclear markers to synthesize emerging patterns of anagenetic speciation on Ulleung Island.

Phylogenetic relationship and genetic differentiation of Populus caspica and Populus alba using cpDNA and ITS noncoding sequences

Populus caspica Bornm. (section Leuce and subsection Albida), one of the most endangered endemic tree species in the Hyrcanian Forest in Iran, has numerous morphological characteristics that are closely similar to Populus alba; to clarify their taxonomic relatedness and genetic differentiation and thus inform conservation strategies, we used the noncoding regions of chloroplast DNA (cpDNA; trnL-F and trnH-psbA) and the internal transcribed spacer (ITS). Leaf samples were collected from six populations across northern Iran. cpDNA and ITS fragments were amplified by universal primers using the PCR technique and directed sequencing. The results showed that P. caspica is genetically differentiated from P. alba, and two ITS variants were detected within some P. caspica individuals. Conflicts between topologies from ITS and plastid genomes were observed. High differentiation of P. caspica from the other Populus species shown in this study confirmed the diverging taxonomic status of this endangered species. We recommend in situ conservation measures (e.g., protected areas) for at least several populations of this species, especially in the plain regions of the Hyrcanian forest.

Genome Size, Molecular Phylogeny, and Evolutionary History of the Tribe Aquilarieae (Thymelaeaceae), the Natural Source of Agarwood.

The tribe Aquilarieae of the family Thymelaeaceae consists of two genera, Aquilaria and Gyrinops, with a total of 30 species, distributed from northeast India, through southeast Asia and the south of China, to Papua New Guinea. They are an important botanical resource for fragrant agarwood, a prized product derived from injured or infected stems of these species. The aim of this study was to estimate the genome size of selected Aquilaria species and comprehend the evolutionary history of Aquilarieae speciation through molecular phylogeny. Five non-coding chloroplast DNA regions and a nuclear region were sequenced from 12 Aquilaria and three Gyrinops species. Phylogenetic trees constructed using combined chloroplast DNA sequences revealed relationships of the studied 15 members in Aquilarieae, while nuclear ribosomal DNA internal transcribed spacer (ITS) sequences showed a paraphyletic relationship between Aquilaria species from Indochina and Malesian. We exposed, for the first time, the estimated divergence time for Aquilarieae speciation, which was speculated to happen during the Miocene Epoch. The ancestral split and biogeographic pattern of studied species were discussed. Results showed no large variation in the 2C-values for the five Aquilaria species (1.35–2.23 pg). Further investigation into the genome size may provide additional information regarding ancestral traits and its evolution history.

Combined Analyses of Chloroplast DNA Haplotypes and Microsatellite Markers Reveal New Insights Into the Origin and Dissemination Route of Cultivated Pears Native to East Asia.

Asian pear plays an important role in the world pear industry, accounting for over 70% of world total production volume. Commercial Asian pear production relies on four major pear cultivar groups, Japanese pear (JP), Chinese white pear (CWP), Chinese sand pear (CSP), and Ussurian pear (UP), but their origins remain controversial. We estimated the genetic diversity levels and structures in a large sample of existing local cultivars to investigate the origins of Asian pears using twenty-five genome-covering nuclear microsatellite (simple sequence repeats, nSSR) markers and two non-coding chloroplast DNA (cpDNA) regions (trnL-trnF and accD-psaI). High levels of genetic diversity were detected for both nSSRs (HE = 0.744) and cpDNAs (Hd = 0.792). The major variation was found within geographic populations of cultivated pear groups, demonstrating a close relationship among cultivar groups. CSPs showed a greater genetic diversity than CWPs and JPs, and lowest levels of genetic differentiation were detected among them. Phylogeographical analyses indicated that the CSP, CWP, and JP were derived from the same progenitor of Pyrus pyrifolia in China. A dissemination route of cultivated P. pyrifolia estimated by approximate Bayesian computation suggested that cultivated P. pyrifolia from the Middle Yangtze River Valley area contributed the major genetic resources to the cultivars, excluding those of southwestern China. Three major genetic groups of cultivated Pyrus pyrifolia were revealed using nSSRs and a Bayesian statistical inference: (a) JPs; (b) cultivars from South-Central China northward to northeastern China, covering the main pear production area in China; (c) cultivars from southwestern China to southeastern China, including Yunnan, Guizhou, Guangdong, Guangxi, and Fujian Provinces. This reflected the synergistic effects of ecogeographical factors and human selection during cultivar spread and improvement. The analyses indicated that UP cultivars might be originated from the interspecific hybridization of wild Pyrus ussuriensis with cultivated Pyrus pyrifolia. The combination of uniparental DNA sequences and nuclear markers give us a better understanding of origins and genetic relationships for Asian pear groups and will be beneficial for the future improvement of Asian pear cultivars.

Phylogeographic analysis suggests two origins for the riparian azalea Rhododendron indicum (L.) Sweet.

Ecological speciation is an important factor in the diversification of plants. The distribution of the woody species Rhododendron indicum, which grows along rivers and is able to withstand water flow when rivers flood (i.e. it is a rheophyte), is disjunct, in contrast to the widespread distribution of its relative, Rhododendron kaempferi. This study aimed to elucidate the phylogenetic relationships between R. indicum and R. kaempferi and the evolutionary processes that gave rise to them. The sequences of three non-coding chloroplast DNA regions (total length 1977 bp) were obtained from 21 populations covering the ranges of the two species. In addition, genome-wide SNPs were genotyped from 20 populations using a genotyping by sequencing method. Leaf morphologies were measured for eight representative populations. Two chloroplast DNA haplotypes, which were detected in R. indicum, were shared between the two species. Genome-wide SNPs identified two lineages in R. indicum and these lineages did not constitute a monophyletic group. Each of these two lineages was related to geographically close populations of R. kaempferi. Leaf morphology, which is a characteristic feature in rheophytes, was not differentiated between the two lineages in R. indicum. The morphological similarity between the two heterogeneous lineages may be a result of parallel evolution from R. kaempferi or of introgressive hybridization between the species due to strong selective pressure imposed by flooding.

Phylogeography and conservation genetics of the rare and relict Bretschneidera sinensis (Akaniaceae).

Bretschneidera sinensis, a class-I protected wild plant in China, is a relic of the ancient Tertiary tropical flora endemic to Asia. However, little is known about its genetics and phylogeography. To elucidate the current phylogeographic patterns and infer the historical population dynamics of B. sinensis, and to make recommendations for its conservation, three non-coding regions of chloroplast DNA (trnQ-rps16, rps8-rps11, and trnT-trnL) were amplified and sequenced across 256 individuals from 23 populations of B. sinensis, spanning 10 provinces of China. We recognized 13 haplotypes, demonstrating relatively high total haplotype diversity (hT = 0.739). Almost all of the variation existed among populations (98.09%, P < 0.001), but that within populations was low (1.91%, P < 0.001). Strong genetic differentiation was detected among populations (GST = 0.855, P < 0.001) with limited estimations of seed flow (Nm = 0.09), indicating that populations were strongly isolated from one another. According to SAMOVA analysis, populations of B. sinensis in China could be divided into five geographic groups: (1) eastern Yunnan to western Guangxi; (2) Guizhou-Hunan-Hubei; (3) central Guangdong; (4) northwestern Guangdong; and (5) the Luoxiao-Nanling-Wuyi -Yangming Mountain. Network analysis showed that the most ancestral haplotypes were located in the first group, i.e., the eastern Yungui Plateau and in eastern Yunnan, which is regarded as a putative glacial refugia for B. sinensis in China. B. sinensis may have expanded its range eastward from these refugia and experienced bottleneck or founder effects in southeastern China. Populations in Liping (Guizhou Province), Longsheng (Guangxi Province), Huizhou (Guangdong Province), Chongyi (Jiangxi Province), Dong-an (Hunan Province), Pingbian (Yunnan Province) and Xinning (Hunan Province) are proposed as the priority protection units.

The Origin of New Natural hybrid, Goodyera × maximo-velutina (Orchidaceae) from Jeju Island, Korea

We describe a new taxon of Goodyera (Orchidaceae), G. × maximo-velutina from Jeju Island. This taxon is presumed to be a natural hybrid between G. maximowicziana and G. velutina based on morphological characteristics, i.e., leaf margin, leaf venation, hairiness and color of floral bract, pedicel and ovary. We analyzed ITS of nuclear DNA and three noncoding regions of chloroplast DNA (trnH-psbA, trnS-trnG, trnL) for elucidating the hybrid origin of the new species, G. × maximo-velutina, and two putative parental species. The morphological characteristics and illustrations of the species based on the holotype and other specimens from natural populations are provided together with photographs of the habitat.