Restriction Site-associated DNA Sequencing Research Articles

Genetic diversity analysis and core collection construction of tea plant from the Yunnan Province of China using ddRAD sequencing

Tea plants are economically important woody plants that originated in southwestern China. The Yunnan Province in China is regarded as the central origin of tea plants owing to the abundance and diversity of the tea germplasm present in this region. However, there is a lack of knowledge regarding the genetic diversity and evolutionary relationships among tea plants in this region. Here, a total of 266,397 high-quality single nucleotide polymorphisms were obtained using double digest restriction-site associated DNA sequencing from 468 tea accessions collected from Yunnan. The phylogenetic relationship and population structure of the 468 tea accessions revealed remarkable inter- and intraspecific introgression across different sect. Thea species. We found that Camellia taliensis (W. W. Smith) Melchior is the main genetic donor that greatly contributed to the domestication of C. sinensis (L.) O. Kuntze, as evidenced by the frequent generation of genetic intermediates derived from the hybridization between C. sinensis and C. taliensis in Lincang, Xishuangbanna, and Pu’er. Notable genetic differences were observed across the populations from distinct regions of Yunnan. Additionally, the Lancang River may have attenuated the genetic interflow between Lincang and Pu’er. Finally, a core collection of 50 tea accessions was constructed based on the genetic diversity of the 468 tea accessions. Our results provide novel insights into the evolutionary and domestication history of tea plants in Yunnan Province and can facilitate the development of strategies for the conservation, breeding, and utilization of the core collection.

Construction of a high-density genetic map and QTL mapping of growth traits in kiwifruit

The trunk diameter (TD) and annual branch diameter (ABD) are closely associated with the growth vigor, abiotic resistance and fruit yield of kiwifruit. These traits are quantitative traits that are controlled by more than one quantitative trait loci (QTLs). However, QTLs for TD and ABD in kiwifruit remain unelucidated. In this study, we generated a cross of the diploid species Actinidia chinensis ‘Hongyang’ (♀) × ‘Boshanbiyu’ (♂) and performed restriction-site associated DNA sequencing on the parents and 173 F1 progeny to identify single-nucleotide polymorphisms (SNPs) in the population. A high-density linkage map spanning 2,548.97 cM was developed using 6,506 bin markers. This bin map consisted of 29 linkage group (LG), with an average genetic distance of 0.39 cM. A total of 8 QTLs were identified: four for TD and four for ABD, with the variance explained by these QTLs ranging from 4.43 % to 16.11 %. A total of 358 TD genes and 181 ABD genes were extracted from the mapped QTLs, and analyzed by KEGG enrichment. In addition, we developed a TD marker from LG23 and an ABD marker from LG10 which were significantly correlated with growth traits. The TD marker is associated with Actinidia22292, a gene involved in protein polar localization, while the ABD marker is linked to Actinidia40478, an Ein3-binding F-box protein. The two SNP markers developed in this study were further validated in a biparental population, indicating their potential use in predicting seedling growth vigor. In summary, these genetic findings deepen our understanding of the genetic basis of growth traits in kiwifruit, and provide valuable information to facilitate kiwifruit breeding.

Divergent Population Structure in Five Common Rockfish Species of Puget Sound, WA Suggests the Need for Species-Specific Management.

Quantifying connectivity between endangered or threatened marine populations is critical information for management and conservation, especially where abundance and productivity differ among such populations. Spatial patterns of such connectivity depend not only on extrinsic factors such as oceanography and bathymetry but also on intrinsic species-specific factors such as life history, demography and the location of glacial refugia. Nevertheless, population structure is often inferred from related or ecologically similar species. For example, the population structure in most rockfish species (Sebastes spp.) in the Salish Sea and the US West Coast is currently inferred from genetic data of three species that are known to hybridise in Puget Sound. Here, we determined the population structure and connectivity in five Puget Sound Rockfish species (Black [Sebastes melanops], Yellowtail [S. flavidus], Redstripe [S. proriger], Greenstriped [S. elongatus], and Puget Sound Rockfish [S. emphaeus]) from over 12,000 restriction-site associated DNA sequencing (RADseq) loci. We found species-specific patterns of genetic differentiation, attributable to both extrinsic and intrinsic factors. Specifically, Black and Puget Sound rockfishes showed no genetic differentiation; Yellowtail and Greenstriped rockfishes were structured according to known geographic barriers; and Redstripe Rockfish revealed evidence for temporal genetic differentiation, suggesting irregular recruitment influences population structure. Only Yellowtail Rockfish followed the federal DPS boundaries generally assumed for rockfish, further emphasizing the importance of species-specific management for the effective recovery and management of these rockfish populations and of marine species in general.

High Crimean-Congo hemorrhagic fever incidence linked to greater genetic diversity and differentiation in Hyalomma marginatum populations in Türkiye

BackgroundTicks are crucial vectors of a wide range of pathogens, posing significant threats to human and animal health globally. Understanding the genetic basis of tick biology and host–parasite interactions is essential for developing effective control programs. This study investigates the fine-scale genetic structure of Hyalomma marginatum Koch, 1844, the primary vector of Crimean-Congo hemorrhagic fever (CCHF) in Türkiye. Despite its significant public health importance, information regarding its population structure and genetic diversity is quite limited.MethodsWe used restriction site-associated DNA sequencing (RAD-Seq) to obtain genome-wide sequence data from 10 tick populations in Türkiye, collected from regions with low, moderate, and high incidence rates of CCHF. Based on these data, we determined population structure and diversity of populations using principal component analysis (PCA) and admixture analysis. Furthermore, we calculated pairwise FST and utilized discriminant analysis of principal components (DAPC) to understand genetic differentiation between populations.ResultsPCA and admixture analysis indicated minimal genetic structure between populations, but we detected notable genetic differentiation and high genetic diversity from regions with high CCHF rates. Furthermore, our DAPC identified 31 significant single-nucleotide polymorphisms (SNPs) associated with regions with high CCHF incidence, with 25 SNPs located near genes involved in critical biological functions such as nucleic acid binding, transmembrane transport, and proteolysis. These findings suggest that genetic variations in these regions may confer adaptive advantages in environments with high pathogen loads.ConclusionsThis study provides the first comprehensive analysis of H. marginatum genetic diversity in Türkiye, revealing significant differentiation in populations from CCHF-endemic regions. These results underscore the importance of considering fine-scale genetic diversity to fully understand the drivers of genetic variation in ticks and their implications for vectorial capacity.Graphical

Population connectivity of the European squid Loligo vulgaris along the West Iberian Peninsula coast: comparing mtDNA and SNPs

Understanding genetic diversity and population connectivity in marine organisms is essential for fisheries management. In the present study, we examined the population genetics of the European squid, Loligo vulgaris, along the western Iberian Peninsula at two genetic resolutions using the mitochondrial cytochrome oxidase subunit I gene (COI) and genomic markers obtained via double digest restriction-site associated DNA sequencing (ddRADseq). The results obtained revealed 79 haplotypes out of 160 COI sequences, while the SNP data set included 86,431 loci after filtering for 38 individuals with 86,319 neutral data. Mitochondrial COI analyses revealed high haplotype (0.961) and nucleotide (0.010) diversities, and the haplotype network reveals complex sub-structure in Turkish waters within a panmictic population. Both Tajima’s D and Fu’s Fs tests suggest that the population of L. vulgaris analysed is evolving neutrally. Pairwise Fst for neutral SNPs were low (0 < Fst<0.002) and not significant showing high homogeneity among populations, while pairwise comparations for candidate adaptive SNPs (112 loci) showed Fst values ranging from 0.026 to 0.234. While neutral SNPs showed admixture, the candidate adaptive SNPs showed a moderate significant structure with a latitudinal discrimination. Overall, both genetic approaches showed homogeneity and strong genetic flux identifying a unique population along the Western Iberian Peninsula.

Morphological and molecular data demonstrate the existence of Betula fruticosa and B. middendorffii and the absence of B. ovalifolia in northeastern China

Abstract Shrub birches are important components of massive wetlands in Northeast China, although much of these habitats have been lost over the past two decades. However, the taxonomy of shrub birches has been ambiguous owing to their morphological variation. In this study, we incorporated morphological and molecular data to address the taxonomic challenges associated with shrub birches. We characterized the morphological variation in the leaves, seeds, and bracts of 11 shrub birch populations in Northeast China and a population of Betula ovalifolia in northern Japan. We genotyped birch individuals from Northeast China at 15 microsatellite loci as well as a subset of individuals, using restriction site-associated DNA sequencing (RAD-seq). The populations in Northeast China consisted of B. fruticosa and B. middendorffii. These two species and B. ovalifolia differ in the width of their seed wings and the angle between the side lobes of their bracts, despite substantial intraspecific variation. Our genetic data revealed two distinct clusters, corresponding to B. fruticosa and B. middendorffii. Ploidy level assessment via RAD-seq revealed that B. fruticosa is diploid and B. middendorffii is tetraploid. Phylogenetic analysis revealed that B. fruticosa formed a sister clade to diploid B. humilis, and B. middendorffii formed a clade with diploid B. nana. Our results indicate the existence of B. fruticosa and B. middendorffii and the probable absence of B. ovalifolia in the northern Daxing’an Range, as all the tetraploid individuals sampled there represented B. middendorffii. In addition, our results suggest that B. humilis and B. nana may have served as diploid parents of the tetraploid species B. ovalifolia and B. middendorffii, respectively.

A pilot study to directly estimate radiation-induced mutation in large Japanese field mouse duo sample, mother and offspring, excluding unknown father, using ddRAD sequencing.

DNA mutations are one of the effects of radiation exposure. A large amount of radioactive materials was released into the environment from the Fukushima Daiichi Nuclear Power Plant after a major earthquake and tsunami. Wild animals and plants living in highly radiation-contaminated areas are constantly exposed to high doses of radiation, and concerns occur about its effects on their health and the next generations. As a pilot study, double-digest restriction site-associated DNA (ddRAD) sequencing was conducted to assess the incidence of mutations in wild large Japanese field mice collected from the evacuation area. The optimal combination of restriction enzymes, encompassing the functionally important coding regions, was selected using in silico analysis. These enzymes were used for ddRAD sequence analysis of females and their fetuses to evaluate mutation rates. The results indicated that no significant differences were observed in mutation rates between mothers and fetuses in the study areas.

Critically ill patients will benefit from single isolated laminar-air-flow wards by improving the environmental microbial composition

BackgroundHospital-associated infection (HAI) is an important issue in intensive care units (ICUs). We still lack direct evidence on whether the ICU patients and/or the medical system can benefit from single isolated laminar-air-flow (LAF) wards.MethodsHigh-touched-surface (HTS) swabs from 5 sites in two kinds of wards with different ventilation systems were longitudinally collected for 16 S rRNA sequencing and Type IIB restriction site-associated DNA sequencing for Microbiome (2bRAD-M). Samples were collected for 3 months. The clinical data of patients admitted to different wards during the sampling time and the whole year were collected and compared.ResultsThe α-diversity of single wards with isolated LAF was significantly higher than open regions without LAF (p<0.01). β-diversity analysis showed differences between different wards and similarities among the same region. We also identified 3 genera attributed to the most difference between the two kinds of wards. 2bRAD-M analysis further revealed community divergence among different HTS sites. There was an overlap between HTS microbiome profiling and the clinically cultivated pathogens of patients with HAI. People in single wards had a better outcome than those in open regions (p<0.05), indicating that single wards had a protective effect for critically ill patients.ConclusionOverall, there was a prominent difference in the microbiome community between single wards and open regions. Single wards had more balanced communities which may lead to better outcomes for patients. For critically ill patients, single ward is recommended when arranging and constructing.

Human Activity Changed the Genetic Pattern of the Orchid Phaius flavus Population

Human activity often has profound effects on plant growth and evolution. Orchids are the most diverse group of flowering plants and are threatened by habitat fragmentation, over-harvesting, and urbanization. A population of Phaius flavus from Beikengding Mount (BM) in the Fujian Province of China was divided into two patches by road construction. This study evaluated its genetic characteristics using restriction site-associated DNA sequencing (RAD-seq) data, more than seven years post-road construction. The purpose of this study was to explore the impact of road construction on the evolution of isolated patches within a population. The analysis revealed that the genetic diversity of patch B was slightly higher than that of patch A in the BM population of P. flavus. Principal component and phylogenetic analyses, genetic structure and genetic differentiation analysis, and bottleneck detection indicated relatively independent genetic differentiation between the two patches. Thus, the construction of the Y013 village road may have influenced different patches of this population on a genetic level. This study provides a case for understanding the impact of specific human activities on plant populations, and then biodiversity conservation. It is conducive to formulating more effective biological protection strategies to mitigate the damage inflicted by human activities on biodiversity.

Genetic diversity insights from population genomics and machine learning tools for Nordic Arctic charr (Salvelinus alpinus) populations

Arctic charr (Salvelinus alpinus) is a salmonid species of high ecological and commercial value in the Holarctic region. Nevertheless, more information is needed about its underlying genetic diversity and population structure in the Nordics, especially regarding farmed populations. High-throughput sequencing was applied in three Arctic charr populations of anadromous or landlocked origin from Finland, Norway and Sweden. More specifically, the animals from the Swedish and Norwegian populations originated from a major egg supplier and producer, respectively. Furthermore, in the case of the Finnish population, the sampled animals originated from the only active conservation program for Arctic charr in the country with a potential interest in farming. Using double-digest restriction site-associated DNA sequencing (ddRAD-seq) on more than 500 fish, over 2000 single nucleotide polymorphisms (SNPs), both in the form of individual SNPs and as read haplotypes, were used to study the genetic diversity and structure of those populations. Genetic diversity metrics were similar between the Norwegian and the Swedish populations. However, substantially lower (40–50 %) genetic diversity was found in the Finnish population. Moreover, considerable genetic differentiation was implied between the studied populations as the mean fixation index (FST) was above 0.1 in all pairwise comparisons. All populations were easily discernible through either principal component analysis (PCA) or discriminant analysis of principal components (DAPC). In addition, unsupervised machine learning models such as K-means, Gaussian and Bayesian Gaussian mixtures were assessed for their ability to detect genetic clusters. A preceding dimensionality reduction step by PCA resulted in all three models, suggesting that the most probable number of clusters was three. Overall, our study affirmed the utility of the developed ddRAD-seq genotyping method and unveiled the genetic structure of the studied populations, both of which could contribute to their more efficient management by captive breeding.

Study of the Genetic Mechanisms of Siberian Stone Pine (Pinus sibirica Du Tour) Adaptation to the Climatic and Pest Outbreak Stresses Using Dendrogenomic Approach.

A joint analysis of dendrochronological and genomic data was performed to identify genetic mechanisms of adaptation and assess the adaptive genetic potential of Siberian stone pine (Pinus sibirica Du Tour) populations. The data obtained are necessary for predicting the effect of climate change and mitigating its negative consequences. Presented are the results of an association analysis of the variation of 84,853 genetic markers (single nucleotide polymorphisms-SNPs) obtained by double digest restriction-site associated DNA sequencing (ddRADseq) and 110 individual phenotypic traits, including dendrophenotypes based on the dynamics of tree-ring widths (TRWs) of 234 individual trees in six natural populations of Siberian stone pine, which have a history of extreme climatic stresses (e.g., droughts) and outbreaks of defoliators (e.g., pine sawfly [Neodiprion sertifer Geoff.]). The genetic structure of studied populations was relatively weak; samples are poorly differentiated and belong to genetically similar populations. Genotype-dendrophenotype associations were analyzed using three different approaches and corresponding models: General Linear Model (GLM), Bayesian Sparse Linear Mixed Model (BSLMM), and Bayesian-information and Linkage-disequilibrium Iteratively Nested Keyway (BLINK), respectively. Thirty SNPs were detected by at least two different approaches, and two SNPs by all three. In addition, three SNPs associated with mean values of recovery dendrophenotype (Rc) averaged across multiple years of climatic stresses were also found by all three methods. The sequences containing these SNPs were annotated using genome annotation of a very closely related species, whitebark pine (P. albicaulis Engelm.). We found that most of the SNPs with supposedly adaptive variation were located in intergenic regions. Three dendrophenotype-associated SNPs were located within the 10 Kbp regions and one in the intron of the genes encoding proteins that play a crucial role in ensuring the integrity of the plant's genetic information, particularly under environmental stress conditions that can induce DNA damage. In addition, we found a correlation of individual heterozygosity with some dendrophenotypes. Heterosis was observed in most of these statistically significant cases; signs of homeostasis were also detected. Although most of the identified SNPs were not assigned to a particular gene, their high polymorphism and association with adaptive traits likely indicate high adaptive potential that can facilitate adaptation of Siberian stone pine populations to the climatic stresses and climate change.

Noninvasive, epigenetic age estimation in an elasmobranch, the cownose ray (Rhinoptera bonasus)

Age data are essential for estimating life history parameters and are thus critical for population assessment, management, and conservation. Traditional vertebrae-based age estimation in elasmobranchs can be costly, time intensive, of low accuracy, and is by necessity lethal. Herein, epigenetic clocks were developed for an elasmobranch, the cownose ray (Rhinoptera bonasus), using aquarium-born individuals (n = 42) with known dates of birth (age range: 7−7,878 days or 0−21 years) and two tissue types (fin clips and whole blood) that can be sampled in a relatively non-invasive manner. Enzymatically-converted restriction site-associated DNA sequencing (ECrad-seq) was used to identify CpG sites that exhibited age-correlated DNA methylation. The epigenetic clocks developed were highly accurate (mean absolute error, MAE, < 0.75 years) and precise (R2 > 0.98). Age-associated CpG sites were identified across tissues, and a multi-tissue clock was also highly accurate (MAE < 1 year) and precise (R2 = 0.97). Using the developed fin clip clock, three wild-caught individuals of unknown age but managed in aquariums for > 22 years were predicted to be 22.10−23.49 years old. Overall, the results have important implications for future epigenetic clock development and noninvasive age estimation in elasmobranchs.

Population genetics of Macrognathus siamensis (Synbranchiformes: Mastacembelidae): Implications for non-migratory fishery resources in the Mekong River basin

The spotted spiny eel, Macrognathus siamensis is an economically important freshwater fish in the Mekong River basin, which is undergoing dramatic biodiversity changes due to anthropogenic impacts. The species is non-migratory, with a moderate larval duration that facilitates downstream genetic connectivity. Restriction site-associated DNA sequence datasets that includes 3736 and 1244 single nucleotide polymorphisms representing putatively neutral loci, from all geographic sites and without the Tachileik site, indicate strong population structure in this sedentary fish, with a significant isolation-by-distance signature. This structure reflects isolation of tributary populations and downstream dispersal of these distinct populations to mix in the Mekong mainstem and contributes to increased genetic diversity in the lower reaches of the Mekong basin. Genetic data indicates that the dispersal of eggs and larvae downstream is not impeded by Khone Falls and that the falls prevent upstream dispersal of eggs and larvae. Despite this downstream connectivity, there is a strong population structure among above- and below-Khone Falls groups. An analysis of outlier loci putatively under selection provides evidence that the difference between connected metapopulations above and below the falls is due to the fundamental differences in environmental regimes. We hypothesize that instead of a physical barrier, the falls simply represents a demarcation between previously recognized upstream limited-floodplain environments and downstream extensive-floodplain environments. These findings emphasize the need to monitor genetic diversity of key sedentary fishery species to assess whether proposed dams along the river could affect genetic diversity downstream. This genetic diversity is particularly important for the resilience of downstream populations because of the many environmental impacts caused by dams, land use, and climate change. Early detection of reduced downstream genetic diversity could trigger remediation programs to ensure resilience and continued fisheries productivity of important non-migratory fish resources.

Development of Simple Sequence Repeat of Monochamus alternatus (Coleoptera: Cerambycidae) Based on Restriction Site-Associated DNA Sequencing.

Monochamus alternatus, a pest posing a serious threat to coniferous species, such as Pinus massoniana, has had devastating effects on pine forests due to its association with Bursaphelenchus xylophilus. The creation of unique simple sequence repeat (SSR) primers for M. alternatus is crucial, as there has been little study of the species' phylogeography. The aim of this study was to identify and create polymorphic SSR primers by sequencing samples of M. alternatus obtained from three different sampling points using the restriction site-associated DNA sequencing (Red-seq) approach. Subsequently, supplementary samples were integrated, and genetic typing was performed utilizing the identified polymorphic primers. Through comprehensive analysis, a total of 95,612 SSR loci were identified. Among these, mononucleotide repeats (51.43%), dinucleotide repeats (28.79%), and trinucleotide repeats (16.74%) predominated among the SSR motif types. Ultimately, 18 pairs of SSR primers were screened out, demonstrating stable amplification and high polymorphism. Genetic typing revealed that the mean number of alleles (Na) for these primer pairs ranged from 3 to 8, observed heterozygosity (Ho) ranged from 0.133 to 0.733, polymorphic information content (PIC) ranged from 0.294 and 0.783, and Shannon's index (I) ranged from 0.590 to 1.802. This study effectively produced 16 pairs of SSR primers that can be applied to different populations of M. alternatus. As a result, important tools for furthering studies on the phylogeography of pine wood nematodes, creating genetic maps, gene mapping, and carrying out in-depth investigations into gene function have been made available.

Population structure of Desmophyllum pertusum found along the United States eastern continental margin

ObjectiveThe connectivity and genetic structuring of populations throughout a region influence a species’ resilience and probability of recovery from anthropogenic impacts. By gaining a comprehensive understanding of population connectivity, more effective management can be prioritized. To assess the connectivity and population genetic structure of a common cold-water coral species, Desmophyllum pertusum (Lophelia pertusa), we performed Restriction-site Associated DNA Sequencing (RADseq) on individuals from nine sites ranging from submarine canyons off New England to the southeastern coast of the United States (SEUS) and the Gulf of Mexico (GOM). Fifty-seven individuals and 3,180 single-nucleotide polymorphisms (SNPs) were used to assess genetic differentiation.ResultsHigh connectivity exists among populations along the SEUS, yet these populations were differentiated from those to the north off New England and in Norfolk Canyon along the North Atlantic coast of the United States, as well as those in the GOM. Interestingly, Norfolk Canyon, located just north of North Carolina, and GOM populations exhibited low levels of genetic differentiation, corroborating previous microsatellite analyses and signifying gene flow between these populations. Increasing sample sizes from existing populations and including additional sampling sites over a larger geographic range would help define potential source populations and reveal fine-scale connectivity patterns among D. pertusum populations.

Genome-Wide Data Uncover Cryptic Diversity With Multiple Reticulation Events in the Balkan-Anatolian Cardamine (Brassicaceae) Species Complex.

Plant species diversity may be considerably underestimated, especially in evolutionarily complex genera and in diversity hotspots that have enabled long-term species persistence and diversification, such as the Balkan Peninsula. Here, we address the topic of underexplored plant diversity and underlying evolutionary and biogeographic processes by investigating the hygrophytic mountain species complex of Cardamine acris s.l. distributed in the Balkans (three subspecies within C. acris) and northwestern Anatolia (C. anatolica). We performed a series of phylogenetic and phylogeographic analyses based on restriction-site associated DNA sequencing (RADseq) and target enrichment (Hyb-Seq) data in combination with habitat suitability modelling. We found C. anatolica as a clade nested within the Balkan C. acris, probably resulting from a founder event, and uncovered three allopatric cryptic lineages within C. acris subsp. acris, allowing us to recognise a total of six entities in this complex. We observed the deepest genetic split within C. acris subsp. acris in the western Balkans, which was at odds with taxonomy and showed no distribution gap. We inferred vicariance as the most likely process for population divergence in the Balkans, accompanied by gene flow between the recognised entities, which was consistent with the modelled habitat suitability dynamics. Furthermore, we discovered several polyploid populations in C. acris, representing both pure intra- and inter-lineage hybrid polyploids, but detected only minor traces of hybridization with related congeners. Overall, our results illustrate that diverse evolutionary processes may influence the history of mountain plant species in the Balkan Peninsula, including vicariance, reticulation, polyploidization and cryptic diversification.

Differentiation of wild accessions within the Panax bipinnatifidus complex using newly developed polymorphic microsatellite markers

Abstract Panax L., renowned as ginseng genus, is a famous medicinal group of family Araliaceae. Within this genus, the taxa of Panax bipinnatifidus complex are mainly distributed in Himalayas and Hengduan Mountain areas. Due to the complex evolutionary history and short-term rapid radiation, the relationships among species within the complex have not been clearly resolved, and the taxa identification is difficult due to the intermediate morphological traits. This study aimed to use the available restriction-site associated DNA sequence data from 29 individuals of P. bipinnatifidus complex to mine high-polymorphic simple sequence repeat (SSR) markers, with the goal of evaluating their utility in taxa identification. Eleven polymorphic SSR loci were ultimately selected and validated through polymerase chain reactions amplifying across 63 individuals of P. bipinnatifidus complex and 13 individuals of three outgroup species. The subsequent genetic diversity analysis uncovered 76 alleles in total, ranging from 5 to 15 per locus. Observed heterozygosity spanned 0.241–0.512, while expected heterozygosity ranged between 0.345 and 0.644. The genetic kinship analysis revealed a sister relationship between Panax zingiberensis and Panax vietnamensis. The analysis result also supported the classification of samples from Hunan and Hubei provinces into a single genetic unit within the P. bipinnatifidus complex. These newly developed SSR markers will facilitate the identification of wild ginseng plants.

The origin and morphological character evolution of the paleotropical woody bamboos.

The woody bamboos (Bambusoideae) exhibit distinctive biological traits within Poaceae, such as highly lignified culms, rapid shoot growth, monocarpic mass flowering and nutlike or fleshy caryopses. Much of the remarkable morphological diversity across the subfamily exists within a single hexaploid clade, the paleotropical woody bamboos (PWB), making it ideal to investigate the factors underlying morphological evolution in woody bamboos. However, the origin and biogeographical history of PWB remain elusive, as does the effect of environmental factors on the evolution of their morphological characters. We generated a robust and time-calibrated phylogeny of PWB using single nucleotide polymorphisms retrieved from optimized double digest restriction site associated DNA sequencing, and explored the evolutionary trends of habit, inflorescence, and caryopsis type in relation to environmental factors including climate, soil, and topography. We inferred that the PWB started to diversify across the Oligocene-Miocene boundary and formed four major clades, that is, Melocanninae, Racemobambosinae s.l. (comprising Dinochloinae, Greslanlinae, Racemobambosinae s.str. and Temburongiinae), Hickeliinae and Bambusinae s.l. (comprising Bambusinae s.str. plus Holttumochloinae). The ancestor of PWB was reconstructed as having erect habit, indeterminate inflorescence and basic caryopsis. The characters including climbing/scrambling habit, determinate inflorescence, and nucoid/bacoid caryopsis have since undergone multiple changes and reversals during the diversification of PWB. The evolution of all three traits was correlated with, and hence likely influenced by, aspects of climate, topography, and soil, with climate factors most strongly correlated with morphological traits, and soil factors least so. However, topography had more influence than climate or soil on the evolution of erect habit, whereas both factors had greater effect on the evolution of bacoid caryopsis than did soil. Our results provide novel insights into morphological diversity and adaptive evolution in bamboos for future ecological and evolutionary research.

Genome-wide analysis tracks the emergence of intraspecific polyploids in Phragmites australis

Polyploidization plays an important role in plant speciation and adaptation. To address the role of polyploidization in grass diversification, we studied Phragmites australis, an invasive species with intraspecific variation in chromosome numbers ranging from 2n = 36 to 144. We utilized a combined analysis of ploidy estimation, phylogeny, population genetics and model simulations to investigate the evolution of P. australis. Using restriction site-associated DNA sequencing (RAD-seq), we conducted a genome-wide analysis of 88 individuals sourced from diverse populations worldwide, revealing the presence of six distinct intraspecific lineages with extensive genetic admixture. Each lineage was characterized by a specific ploidy level, predominantly tetraploid or octoploid, indicative of multiple independent polyploidization events. The population size of each lineage has declined moderately in history while remaining large, except for the North American native and the US Land types, which experienced constant population size contraction throughout their history. Our investigation did not identify direct association between polyploidization events and grass invasions. Nonetheless, we observed octoploid and hexaploid lineages at contact zones in Romania, Hungary, and South Africa, suggestively due to genomic conflicts arising from allotetraploid parental lineages.

The Range‐Wide Genetic Structure of a High‐Elevation Conifer Species, Subalpine Larch

ABSTRACTAimSubalpine larch is a long‐lived conifer with a restricted distribution at treeline in the mountains of western North America. Climate change is reducing the availability of this habitat and subalpine larch is unlikely to adapt to its changing environment due to its long generation time, relatively low levels of standing genetic variation and the high degree of habitat fragmentation across the species' range. This study seeks to elucidate the genetic structure of subalpine larch to inform future conservation and management efforts.LocationWestern North America.TaxonSubalpine larch (Larix lyallii Parl.).MethodsTissue was collected from high‐elevation populations across the entire species range. Restriction site associated DNA sequencing was used to generate single nucleotide polymorphism (SNP) data. Population genetic analyses identified genetic differentiation.ResultsClustering analyses performed using 751 neutral SNPs identified three genetically differentiated regions: the Cascade Range, the southern Rocky Mountains and the northern Rocky Mountains. AMOVA confirmed significant genetic differentiation among regions. A discriminant analysis of principal components and a dendrogram of Provesti's genetic distance both supported the hypothesis that genetically distinct lineages arose after dispersal from a single Pleistocene refugium. Significant isolation by distance (IBD) supported the key role of dispersal in shaping modern patterns of genetic variation in subalpine larch.Main ConclusionsPost‐Pleistocene dispersal resulted in genetic differentiation among groups of subalpine larch in the Cascade Range, the southern Rocky Mountains and the northern Rocky Mountains. These three regions should be prioritised for future management.