Didymodonchangbaiensis (Pottiaceae, Musci), a new species from Changbai Mountain, China and its phylogenetic position based on molecular data.

Changbai Mountain, located in northeast China, is one of the areas with the most complete natural ecosystem preservation in China. A new species, Didymodonchangbaiensis C.Feng, J.Kou, H.-X. Xiao & T.-T.Wu from north slope of Changbai Mountain in Jilin Province of China is described and illustrated. It is characterised by ovate or ovate-lanceolate leaves that are appressed when dry, acute leaf apex, lamina red or reddish-orange with KOH, unistratose lamina throughout, plane and unistratose leaf margins, percurrent costa with one layer of guide cells and without ventral stereids, upper and middle laminal cells with elliptical papillae over the transverse walls between two immediately adjacent cells and basal laminal cells not differentiated from the median cells. Our morphological analyses and molecular results, based on DNA sequences of ITS, rps4 and trnM-trnV, confirm that D.changbaiensis is revealed to be sister to D.daqingii J. Kou, R.H. Zander & C. Feng. This new species is compared with similar species and its phylogenetic position and ecology are discussed.

Optimization of SPME-GC-MS and characterization of floral scents from Aquilegia japonica and A. amurensis flowers.

BACKGROUND: The floral scents of plants play a key role in plant reproduction through the communication between plants and pollinators. Aquilegia as a model species for studying evolution, however, there have been few studies on the floral scents and relationships between floral scents and pollination for Aquilegia taxa. METHODS: In this study, three types of solid-phase micro-extraction (SPME) fiber coatings (DVB/PDMS, CAR/PDMS, DVB/CAR/PDMS) were evaluated for their performance in extracting volatile organic compounds (VOCs) from flowers of Aquilegia amurensis, which can contribute to the future studies of elucidating the role of floral scents in the pollination process. RESULTS: In total, 55 VOCs were identified, and among them, 50, 47 and 45 VOCs were extracted by the DVB/CAR/PDMS fiber, CAR/PDMS fiber and DVB/PDMS fibers, respectively. Only 30 VOCs were detected in A. japonica taxa. Furthermore, the relative contents of 8 VOCs were significant different (VIP > 1 and p < 0.05) between the A. amurensis and A. japonica. CONCLUSIONS: The results can be applied in new studies of the relationships between the chemical composition of floral scents and the processes of attraction of pollinator. It may provide new ideas for rapid evolution and frequent interspecific hybridization of Aquilegia.

Population genetic structures of two ecologically distinct species Betula platyphylla and B. ermanii inferred based on nuclear and chloroplast DNA markers.

Climatic oscillations during the last glacial maximum (LGM) significantly affected the distribution patterns and genetic structure of extant plants. Northeast China (NEC) is a major biodiversity center in East Asia, and the influence of historical climate change on NEC populations is critical for understanding species responses to future climate change. However, only a few phylogeographic studies of cool temperate deciduous tree species have been conducted in the area, and results are inconsistent for species with different niches or distribution areas. We employed multiple chloroplast and nuclear markers to investigate the genetic structure of two ecologically contrasting species, Betula platyphylla and B. ermanii, in NEC. Rare haplotypes were identified in the chloroplast genome of these species, and both exhibited high levels of nucleotide diversity based on a fragment of the nuclear gene G3PDH and microsatellites. Moreover, significant phylogeographic structure was detected for B. platyphylla, suggesting that these populations had recolonized from independent glacial refuges, whereas no genetic structure was found for B. ermanii. OPEN RESEARCH BADGES: The nSSR datasets used in the current study and the table of pairwise FST (below diagonal) and its standardized F'ST (above diagonal) among 25 populations based on seven SSRs are available from the Dryad (DOI: https://doi.org/10.5061/dryad.230d176). Sequences generated from this study were deposited in GenBank under Accession nos. KY199568-KY200162 and MK819541-MK819970.

Rapid Divergence Followed by Adaptation to Contrasting Ecological Niches of Two Closely Related Columbine Species Aquilegia japonica and A. oxysepala.

Elucidating the mechanisms underlying the genetic divergence between closely related species is crucial to understanding the origin and evolution of biodiversity. The genus Aquilegia L. has undergone rapid adaptive radiation, generating about 70 well-recognized species that are specialized to distinct habitats and pollinators. In this study, to address the underlying evolutionary mechanisms that drive the genetic divergence, we analyzed the whole genomes of two ecologically isolated Aquilegia species, A. oxysepala and A. japonica as well as their putative hybrid. Our comparative genomic analyses reveal that while the two species diverged only recently and experienced recurrent gene flow, a high level of genetic divergence is observed in their nuclear genomes. In particular, candidate genomic regions that show signature of selection differ dramatically between the two species. Given that the splitting time of the two species is broadly matched with the decrease in effective population sizes, we propose that allopatric isolation together with natural selection have preceded the interspecific gene flow in the process of speciation. The observed high genetic divergence is likely an outcome of combined effects of natural selection, genetic drift and divergent sorting of ancestral polymorphisms. Our study provides a genome-wide view of how genetic divergence has evolved between closely related species.

Development of 15 microsatellite markers in Acer triflorum (Aceraceae) and cross-amplification in congeneric species.

PREMISE OF THE STUDY: Acer (Aceraceae) is an important genus in forest ecosystems in the Northern Hemisphere. In China, 151 species have been reported, and approximately 61 species are endemic. Thus, China is considered to host the greatest diversity of Acer, but markers are needed to evaluate the genetic structure and genetic diversity of these populations of wild Acer species. METHODS AND RESULTS: Using an enriched genomic library, we developed and characterized 15 microsatellite primers for A. triflorum, 10 of which were polymorphic. The number of alleles varied from one to nine. The levels of observed heterozygosity and expected heterozygosity per locus ranged from 0.000 to 1.000 and 0.000 to 0.826, respectively. Most primers also successfully amplified in A. ginnala, A. griseum, A. mandshuricum, A. pseudosieboldianum, A. sinopurpurascens, A. tegmentosum, and A. ukurunduense. CONCLUSIONS: These markers from A. triflorum will provide an opportunity to study genetic diversity and genetic structure in the genus Acer.

Positive Selection Driving Cytoplasmic Genome Evolution of the Medicinally Important Ginseng Plant Genus Panax.

Panax L. (the ginseng genus) is a shade-demanding group within the family Araliaceae and all of its species are of crucial significance in traditional Chinese medicine. Phylogenetic and biogeographic analyses demonstrated that two rounds of whole genome duplications accompanying with geographic and ecological isolations promoted the diversification of Panax species. However, contributions of the cytoplasmic genomes to the adaptive evolution of Panax species remained largely uninvestigated. In this study, we sequenced the chloroplast and mitochondrial genomes of 11 accessions belonging to seven Panax species. Our results show that heterogeneity in nucleotide substitution rate is abundant in both of the two cytoplasmic genomes, with the mitochondrial genome possessing more variants at the total level but the chloroplast showing higher sequence polymorphisms at the genic regions. Genome-wide scanning of positive selection identified five and 12 genes from the chloroplast and mitochondrial genomes, respectively. Functional analyses further revealed that these selected genes play important roles in plant development, cellular metabolism and adaptation. We therefore conclude that positive selection might be one of the potential evolutionary forces that shaped nucleotide variation pattern of these Panax species. In particular, the mitochondrial genes evolved under stronger selective pressure compared to the chloroplast genes.

The impacts of polyploidy, geographic and ecological isolations on the diversification of Panax (Araliaceae).

BACKGROUND: Panax L. is a medicinally important genus within family Araliaceae, where almost all species are of cultural significance for traditional Chinese medicine. Previous studies suggested two independent origins of the East Asia and North America disjunct distribution of this genus and multiple rounds of whole genome duplications (WGDs) might have occurred during the evolutionary process. RESULTS: We employed multiple chloroplast and nuclear markers to investigate the evolution and diversification of Panax. Our phylogenetic analyses confirmed previous observations of the independent origins of disjunct distribution and both ancient and recent WGDs have occurred within Panax. The estimations of divergence time implied that the ancient WGD might have occurred before the establishment of Panax. Thereafter, at least two independent recent WGD events have occurred within Panax, one of which has led to the formation of three geographically isolated tetraploid species P. ginseng, P. japonicus and P. quinquefolius. Population genetic analyses showed that the diploid species P. notoginseng harbored significantly lower nucleotide diversity than those of the two tetraploid species P. ginseng and P. quinquefolius and the three species showed distinct nucleotide variation patterns at exon regions. CONCLUSION: Our findings based on the phylogenetic and population genetic analyses, coupled with the species distribution patterns of Panax, suggested that the two rounds of WGD along with the geographic and ecological isolations might have together contributed to the evolution and diversification of this genus.

Genetic and Epigenetic Diversities Shed Light on Domestication of Cultivated Ginseng (Panax ginseng).

Chinese ginseng (Panax ginseng) is a medically important herb within Panax and has crucial cultural values in East Asia. As the symbol of traditional Chinese medicine, Chinese ginseng has been used as a herbal remedy to restore stamina and capacity in East Asia for thousands of years. To address the evolutionary origin and domestication history of cultivated ginseng, we employed multiple molecular approaches to investigate the genetic structures of cultivated and wild ginseng across their distribution ranges in northeastern Asia. Phylogenetic and population genetic analyses revealed that the four cultivated ginseng landraces, COMMON, BIANTIAO, SHIZHU, and GAOLI (also known as Korean ginseng), were not domesticated independently and Fusong Town is likely one of the primary domestication centers. In addition, our results from population genetic and epigenetic analyses demonstrated that cultivated ginseng maintained high levels of genetic and epigenetic diversity, but showed distinct cytosine methylation patterns compared with wild ginseng. The patterns of genetic and epigenetic variation revealed by this study have shed light on the domestication history of cultivated ginseng, which may serve as a framework for future genetic improvements.

Phenotypic and genetic evidence for ecological speciation of Aquilegia japonica and A. oxysepala.

Natural selection is thought to be a driving force that can cause the evolution of reproductive isolation. The genus Aquilegia is a model system to address how natural selection promotes the process of speciation. Morphological differences between A. oxysepala, A. japonica and their hybrids were quantified for two vegetative (plant height and leaf area) and three floral morphological (sepal area, corolla length and diameter) traits. We also evaluated the genetic variability of the two species and their hybrids based on two chloroplast (1225 bp), four nuclear (5811 bp) genes and 15 microsatellites. Our results revealed that differentiation of A. japonica and A. oxysepala at the ecological and morphological levels also involved divergence at the genetic level. In addition, the analysis of nucleotide variation patterns showed that the two species possessed numerous fixation sites at nuclear genes gAA4, gA7 and gAA12. Furthermore, we found that all of the phenotypic hybrids also showed a genetically admixed ancestry. These findings suggest that natural selection has indeed facilitated the formation of distinct genetic variation patterns in the two Aquilegia species and habitat adaptation has been driving the ecologically based evolution of reproductive isolation.

A simple strategy for development of single nucleotide polymorphisms from non-model species and its application in Panax.

Single nucleotide polymorphisms (SNPs) are widely employed in the studies of population genetics, molecular breeding and conservation genetics. In this study, we explored a simple route to develop SNPs from non-model species based on screening the library of single copy nuclear genes (SCNGs). Through application of this strategy in Panax, we identified 160 and 171 SNPs from P. quinquefolium and P. ginseng, respectively. Our results demonstrated that both P. ginseng and P. quinquefolium possessed a high level of nucleotide diversity. The number of haplotype per locus ranged from 1 to 12 for P. ginseng and from 1 to 9 for P. quinquefolium, respectively. The nucleotide diversity of total sites (πT) varied between 0.000 and 0.023 for P. ginseng and 0.000 and 0.035 for P. quinquefolium, respectively. These findings suggested that this approach is well suited for SNP discovery in non-model organisms and is easily employed in standard genetics laboratory studies.

Isolation and identification of fourteen microsatellite markers in Clivia miniata and Clivia nobilis (Amaryllidaceae).

Clivia is a genus of great horticultural importance and has been widely cultivated as ornamental plants in all over the world. In order to assess the phylogenetic relationships and genetic diversity of the wild Clivia species and cultivars, we isolated AC-enriched repeats using FIASCO from a single clone each of C. miniata Regel. and Clivia nobilis Lindl. Of the fourteen repeats, 10 were polymorphic and 4 were monomorphic. The polymorphic marker loci were characterized using 61 Clivia accessions. The number of alleles ranged from two to six, observed heterozygosity ranged from 0.04 to 1.00 and expected heterozygosity ranged from 0.04 to 0.83. These microsatellite marker loci provide tools for future studies of Clivia species and cultivars.

Genomic and EST-derived microsatellite markers for Iris laevigata (Iridaceae) and other congeneric species.

PREMISE OF THE STUDY: The aims of this study are to develop and characterize genomic and expressed sequence tag (EST)-derived microsatellites from Iris laevigata and test their transferability in I. ensata, I. setosa, I. halophila, I. scariosa, I. potaninii, I. tenuifolia, I. bloudowii, and I. sanguinea. METHODS AND RESULTS: Ten genomic and six EST-derived microsatellites were characterized in I. laevigata. These microsatellite primers amplified one to five alleles in I. laevigata and some of these primers were also successfully amplified in congeneric species. CONCLUSIONS: These microsatellite primers provide us an initial set of molecular markers to explore the spatial population genetic structure of I. laevigata. In addition, these markers may also be useful in population and conservation genetic studies of closely related species.

Genomic and EST microsatellite markers for Aquilegia flabellata and cross-amplification in A. oxysepala (Ranunculaceae).

PREMISE OF THE STUDY: The goal of this study was to identify genomic and expressed sequence tag (EST)-derived microsatellite markers from the species Aquilegia flabellata and assess their transferability in A. oxysepala. METHODS AND RESULTS: Eleven genomic and nine EST-derived microsatellite loci were characterized in A. flabellata. These microsatellite primers amplified 66 alleles in all 20 loci. The observed heterozygosity (H(O)) for each population ranged from 0.00 to 0.94. CONCLUSIONS: All of the 20 loci were successfully amplified in A. oxysepala. These genomic and EST-derived microsatellite markers will be useful in further genetic structure, speciation, and adaptive evolution studies.

Genomic and EST microsatellites for Rhododendron aureum (Ericaceae) and cross-amplification in other congeneric species.

PREMISE OF THE STUDY: Our objective of this study was to develop genomic and expressed sequence tag (EST)-derived microsatellites for the endangered species Rhododendron aureum and access their transferability in R. dauricum and R. brachycarpum. • METHODS AND RESULTS: Twelve genomic microsatellites were isolated in R. aureum using the Fast Isolation by AFLP of Sequences Containing repeats (FIASCO) protocol and seven EST-derived microsatellites were characterized by screening the Rhododendron dbEST database of GenBank. The number of alleles per locus ranged from one to eight. The observed and expected heterozygosity varied from 0.00 to 1.00 and from 0.00 to 0.73, respectively. • CONCLUSIONS: A total of 19 microsatellite loci were developed for R. aureum. Sixteen and 10 of these loci were successfully amplified in R. brachycarpum and R. dauricum, respectively. These microsatellite markers will have potential applications in genetic diversity and conservation genetics studies.