Non-coding Chloroplast DNA Research Articles

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.

Diversity among rare and common congeneric plant species from the Garry oak and Okanagan shrub-steppe ecosystems in British Columbia: implications for conservation

It is often assumed that the northern peripheral populations of species’ ranges are genetically depauperate due in part to founder effects from postglacial colonization. The majority of federally protected plant species are peripheral in Canada, yet we have little information about their patterns of genetic diversity and structure. In British Columbia, the majority of these protected plant species occur in two threatened habitats: the Garry oak and Okanagan shrub-steppe ecosystems. Using universal noncoding chloroplast DNA markers, we investigated genetic diversity and genetic structure in four rare and common plant species pairs inhabiting these two ecosystems. We found that rare species had lower genetic diversity than their common congeners, and detected contrasting patterns of regional diversity and structure based on ecosystem. Species from the Garry oak ecosystem showed lower genetic diversity in the northern deglaciated region and significant differentiation between regions, likely due to limited dispersal between Vancouver Island and the mainland. Species from the Okanagan shrub-steppe, however, tended to have uniform diversity across their range and lack regional structure. This study provides an important first look at the phylogeographic patterns of four rare plant species in British Columbia.

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.

Substitution rate heterogeneity across hexanucleotide contexts in noncoding chloroplast DNA.

Substitutions between closely related noncoding chloroplast DNA sequences are studied with respect to the composition of the 3 bases on each side of the substitution, that is the hexanucleotide context. There is about 100-fold variation in rate, among the contexts, particularly on substitutions of A and T. Rate heterogeneity of transitions differs from that of transversions, resulting in a more than 200-fold variation in the transitions: transversion bias. The data are consistent with a CpG effect, and it is shown that both the A + T content and the arrangement of purines/pyrimidines along the same DNA strand are correlated with rate variation. Expected equilibrium A + T content ranges from 36.4% to 82.8% across contexts, while G–C skew ranges from −77.4 to 72.2 and A–T skew ranges from −63.9 to 68.2. The predicted equilibria are associated with specific features of the content of the hexanucleotide context, and also show close agreement with the observed context-dependent compositions. Finally, by controlling for the content of nucleotides closer to the substitution site, it is shown that both the third and fourth nucleotide removed on each side of the substitution directly influence substitution dynamics at that site. Overall, the results demonstrate that noncoding sites in different contexts are evolving along very different evolutionary trajectories and that substitution dynamics are far more complex than typically assumed. This has important implications for a number of types of sequence analysis, particularly analyses of natural selection, and the context-dependent substitution matrices developed here can be applied in future analyses.

Juniperus excelsa s. str. in crimea – differentiation and history inferred from genetic and morphological markers

Abstract Juniperus excelsa s. str. (Greek juniper) in Crimea is a relic species on the limits of its range, and represents the Mediterranean flora in the Sub-Mediterranean part of the peninsula. Its origin and history in this area remains unresolved. We aimed to analyze phylogeny and potential demographic expansion of the juniper in the Crimea as well as to study its morphological differentiation. We analyzed plant material from 59 trees inhabiting eight populations. Genetic variation assessments were based on the four non-coding chloroplast DNA (cpDNA) fragments and the nuclear internal transcribed spacer region ITS1-5,8S-ITS2 (ITS). To examine the morphological differentiation, eight measured/counted traits of cones, seeds, and shoots were chosen and eight ratios were calculated. Morphological parameters were compared using ANOVA, Student’s t test, discrimination analysis and Kruskal-Wallis and U Mann-Whitney tests. Two cpDNA fragments were polymorphic and, in total, 10 cpDNA haplotypes were found. Haplotype diversity (Hd) ranged from 0.0 to 0.9. Based on both cpDNA and ITS sequences variation, phylogenetic analyses revealed a close relationship of the Crimean junipers to the individuals from other parts of the species range. In general, our molecular results confirmed the low level of genetic differentiation of J. excelsa individuals inhabiting different parts of the species range, likely resulting from a common ancestry. Only slight morphological differences were found between populations with different geographic location or habitat. The analyzes showed the distinctness of the populations from the southern part of the coast. Some unique morphological and molecular features of southern coastal populations imply that they are remnants of Late Pleistocene abundant forests. We suggest that the recent fragmentation of the Juniperus populations in the Crimean Peninsula could have arisen during the Atlantic period of the Holocene.

The transition to agricultural cultivation of neo-crops may fail to account for wild genetic diversity patterns: insights from the Cape Floristic Region.

AimThe global increase in the cultivation of native wild plants has raised concerns regarding potential risks associated with translocating genetic lineages beyond their natural range. This study aimed to investigate whether agricultural cultivation of neo-crops (a) accounts for the levels of genetic diversity present in wild populations, and whether (b) cultivated populations are genetically divergent from wild populations and thus pose a potential threat to wild genetic diversity.LocationThe Cape Floristic Region (CFR), located along the southern Cape of South Africa.MethodsHigh Resolution Melt analysis (HRM) coupled with Sanger sequencing was used to screen three non-coding chloroplast DNA loci in Cyclopia Vent. (Fabaceae), a CFR endemic neo-crop cultivated for the production of a herbal infusion referred to as Honeybush tea. Wild and cultivated populations for three of three widely cultivated Honeybush species (C. intermedia, C. longifolia, and C. subternata) were screened. Genetic diversity and differentiation were measured and compared between wild and cultivated groups.ResultsAcross all asseccions, a total of 17 haplotypes were detected, four of which were shared between wild and cultivated populations, while the remaining 13 were only detected in wild populations. Genetic diversity and differentiation was significantly higher in wild populations than in cultivated populations.ConclusionsIf no guidelines exist to facilitate the introduction of native wild plant taxa to a cultivated setting, wild genetic diversity patterns are likely to be compromised by cultivated populations. In the case presented here, cultivation represents a genetic bottleneck, failing to account for rare haplotypes, and may have disrupted species boundaries by initiating interspecific hybridization. More empirical work is required to evaluate the extent to which neo-crop cultivation poses a risk to wild genetic resources in the CFR and globally.

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.

Aegilops tauschii Coss. molecular phylogeny: nuclear gene Got2 versus chloroplast DNA data

Aegilops tauschii Coss. phylogenetic tree based on DNA sequences of nuclear gene Got2, about 3000 b.p., studied in 114 accessions of the species was constructed and compared with the tree previously built using non-coding chloroplast DNA (cpDNA), about 3000 b.p. in total, in 111 Ae. tauschii accessions. (110 accessions are the same on both trees.) The two trees differ greatly, implying that despite of Ae. tauschii being mostly self-pollinating species, cross pollination played an important role in the species evolutionary history. Got2 tree rather well corresponds to Ae. tauschii intraspecies systematics based on spike morphology, while cpDNA tree is in contradiction with this systematics. Ae. tauschii phylogeographic patterns essentially differ in Ae. tauschii subsp. tauschii and Ae. tauschii subsp. strangulata. As revealed by Got2 DNA data, subsp. strangulata presents five major common phylogenetic lineages sharing the subspecies range between each other, and minor relict lineages in addition. Subspecies tauschii presents single major lineage occupying all the subspecies range, and additional relict lineages.

Phylogeographic and demographic modeling analyses of the multiple origins of the rheophytic goldenrod Solidago yokusaiana Makino.

Understanding adaptation mechanisms is important in evolutionary biology. Parallel adaptation provides good opportunities to investigate adaptive evolution. To confirm parallel adaptation, it is effective to examine whether the phenotypic similarity has one or multiple origins and to use demographic modeling to consider the gene flow between ecotypes. Solidago yokusaiana is a rheophyte endemic to the Japanese Archipelago that diverged from Solidago virgaurea. This study examined the parallel origins of S. yokusaiana by distinguishing between multiple and single origins and subsequent gene flow. The haplotypes of noncoding chloroplast DNA and genotypes at 14 nuclear simple sequence repeat (nSSR) loci and single-nucleotide polymorphisms (SNPs) revealed by double-digest restriction-associated DNA sequencing (ddRADseq) were used for phylogeographic analysis; the SNPs were also used to model population demographics. Some chloroplast haplotypes were common to S. yokusaiana and its ancestor S. virgaurea. Also, the population genetic structures revealed by nSSR and SNPs did not correspond to the taxonomic species. The demographic modeling supported the multiple origins of S. yokusaiana in at least four districts and rejected a single origin with ongoing gene flow between the two species, implying that S. yokusaiana independently and repeatedly adapted to frequently flooding riversides.

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.

Application of High Resolution Melt analysis (HRM) for screening haplotype variation in a non-model plant genus: Cyclopia (Honeybush)

AimThis study has three broad aims: to (a) develop genus-specific primers for High Resolution Melt analysis (HRM) of members of Cyclopia Vent., (b) test the haplotype discrimination of HRM compared to Sanger sequencing, and (c) provide an example of using HRM to detect novel haplotype variation in wild C. subternata Vogel. populations.LocationThe Cape Floristic Region (CFR), located along the southern Cape of South Africa.MethodsPolymorphic loci were detected through a screening process of sequencing 12 non-coding chloroplast DNA segments across 14 Cyclopia species. Twelve genus-specific primer combinations were designed around variable cpDNA loci, four of which failed to amplify under PCR; the eight remaining were applied to test the specificity, sensitivity and accuracy of HRM. The three top performing HRM Primer combinations were then applied to detect novel haplotypes in wild C. subternata populations, and phylogeographic patterns of C. subternata were explored.ResultsWe present a framework for applying HRM to non-model systems. HRM accuracy varied across the PCR products screened using the genus-specific primers developed, ranging between 56 and 100%. The nucleotide variation failing to produce distinct melt curves is discussed. The top three performing regions, having 100% specificity (i.e. different haplotypes were never grouped into the same cluster, no false negatives), were able to detect novel haplotypes in wild C. subternata populations with high accuracy (96%). Sensitivity below 100% (i.e. a single haplotype being clustered into multiple unique groups during HRM curve analysis, false positives) was resolved through sequence confirmation of each cluster resulting in a final accuracy of 100%. Phylogeographic analyses revealed that wild C. subternata populations tend to exhibit phylogeographic structuring across mountain ranges (accounting for 73.8% of genetic variation base on an AMOVA), and genetic differentiation between populations increases with distance (p < 0.05 for IBD analyses).ConclusionsAfter screening for regions with high HRM clustering specificity—akin to the screening process associated with most PCR based markers—the technology was found to be a high throughput tool for detecting genetic variation in non-model plants.

Phylogeography of the rare and endangered lycophyte Isoetes yunguiensis.

Background Isoetes yunguiensis Q. F. Wang & W. C. Taylor is a lycophyte of an ancient genus, and it is endemic to China. It is a first-class protected plant in China. This living fossil is used in paleoecology and studies on the evolution of Lycophytes in the Yunnan-Guizhou Plateau. In recent years, human activities have caused the disappearance of several wild populations, and the number of plants in the existing populations is low. Study of the genetic structure, distribution pattern, and historical dynamics of I. yunguiensis in all areas of its distribution is of guiding significance for its rational and effective protection.MethodsExpressed sequence tag-simple sequence repeat (EST-SSR) markers were used to study the genetic diversity and structure of I. yunguiensis, and noncoding chloroplast DNA (cpDNA) sequences were used to study the pedigree, population dynamics history, and glacial shelter of I. yunguiensis. A maximum entropy model was used to predict the past, present, and future distribution patterns of I. yunguiensis.ResultsAnalysis with EST-SSR markers revealed that I. yunguiensis showed high genetic diversity and that genetic variation was significantly higher within populations than between populations. Based on cpDNA data, it was concluded that there was no significant geographic pedigree in the whole area of I. yunguiensis distribution (NST = 0.344 > GST = 0.183, p > 0.05); 21 haplotypes were detected using DnaSP v5. Neutral test and LAMARC simulation showed that I. yunguiensis has experienced rapid expansion in recent years. The maximum entropy model predicted that the potential distribution area of I. yunguiensis in the last glacial maximum period has increased significantly compared with the present distribution area, but the future distribution area did not show substantial changes.

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.

Phylogenetic relationship in Actinidia (Actinidiaceae) based on four noncoding chloroplast DNA sequences

The interspecies relationships within Actinidia remain controversial due to lack of sufficient taxon sampling and molecular markers. Intergenic spacers (IGS) in the chloroplast genome have been shown to yield substantial variability and were successfully used in phylogenetic study in diverse plants. Hence, in this study, we develop IGS markers by comparing the available chloroplast genomes of Actinidia to explore the interspecific relationships within Actinidia. A total of 46 species were sampled, and three phylogenetic analyses methods (MP, ML and Bayesian) were used. Compared to the plastomes of other congeneric species, twenty-four highly variable intergenic spacers are identified, and four intergenic spacers with high polymorphism [ndhF-rps32, ndhC-trnV (UAC), rps16-trnQ (UUG) and trnE (UUC)-trnT (GGU)] were used to reconstruct phylogenetic relationships within Actinidia. The phylogeny reveals two main groups within Actinidia, and the four traditionally recognized sections are not monophyletic. Two sections (Leiocarpae and Maculatae) can be proposed, among which Leiocarpae is the earliest diverged lineage in the genus. As compared to previous work, our study provides a robust phylogenetic hypothesis that aids in our understanding of the interspecific relationships of Actinidia. Several closely related species are placed in disparate parts of the phylogeny. Moreover, the same species sampled at least two varieties are polyphyletic in our cpDNA trees, which suggests frequent hybridization/introgression events occurred in the genus of Actinidia.

Chloroplast DNA variation in a hyperdiverse tropical tree community.

We investigate chloroplast DNA variation in a hyperdiverse community of tropical rainforest trees in French Guiana, focusing on patterns of intraspecific and interspecific variation. We test whether a species genetic diversity is higher when it has congeners in the community with which it can exchange genes and if shared haplotypes are more frequent in genetically diverse species, as expected in the presence of introgression.We sampled a total of 1,681 individual trees from 472 species corresponding to 198 genera and sequenced them at a noncoding chloroplast DNA fragment.Polymorphism was more frequent in species that have congeneric species in the study site than in those without congeners (30% vs. 12%). Moreover, more chloroplast haplotypes were shared with congeners in polymorphic species than in monomorphic ones (44% vs. 28%).Despite large heterogeneities caused by genus‐specific behaviors in patterns of hybridization, these results suggest that the higher polymorphism in the presence of congeners is caused by local introgression rather than by incomplete lineage sorting. Our findings suggest that introgression has the potential to drive intraspecific genetic diversity in species‐rich tropical forests.

Aegilops tauschii Coss. chloroplast genome phylogeny

The chloroplast genome phylogeny of Aegilops tauschii was studied. The sequences of four regions of non-coding chloroplast DNA (cpDNA), about 3000 b.p. in total, in 111 Ae. tauschii accessions, 55 of subspecies tauschii (TauL1) and 56 of subspecies strangulata (2 of TauL3 and 54 of TauL2 gene-pool), were used for neighboring joint and maximum likelihood analysis. This revealed that the patterns of cpDNA divergence essentially differ from those of nuclear genome in Ae. tauschii, coinciding only at TauL3, indicating that the chloroplast genome of TauL3 Ae. tauschii differentiated from those of other Ae. tauschii in ancient times. Among the other Ae. tauschii, Taul1 plants have relatively “older” variants of chloroplast genome; moreover, from 55 TauL1 accessions studied, the four accessions originating from the edges of the species area retained the relict variant of cpDNA, with no indels or base pair substitution having taken place in about 3000 b.p. studied, since the differentiation of TauL1 and TauL2 started. Most of TauL1 plants form a cluster with rather similar cpDNA, which also included eight of 54 Taul2 accessions studied, all originating from Caucasia and pre-Caspian Iran. The other 46 TauL2 accessions form a single cluster residing on the tree among the major and several minor, relict, lineages of TauL1. This cluster presents relatively “young”, considerably changed by mutations, and also highly variable cpDNA. It further falls in the two sub-clusters: one presents TauL2 Ae. tauschii from South Caucasia and pre-Caspian Iran, and another—with TauL2 from Caucasia, Continental Iran and Turkmenistan.

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.