Study on Combustion and Emissions of a Combined Injection Spark Ignition Engine with Natural Gas Direct Injection Plus Ethanol Port Injection under Lean-Burn Conditions.

Conventional ethanol spark ignition (SI) engines have poor fuel atomization and mixture formation. The objective of this paper is to improve the combustion and emission performance of ethanol SI engines under lean-burn conditions through the dual-injection mode with ethanol port injection and compressed natural gas (CNG) direct injection (CDI+EPI). This paper studies the engine performance at 1500 rpm under five CNG direct injection ratios (CDIr) and five excess air ratios (λ). The results show that as the CDIr increases under lean-burn conditions, the following occurs: the minimum advance for best torque (MBT), the coefficient of variation (CoVIMEP), and CO and HC emissions decrease; the crankshaft rotation or time with cumulative heat release rate ranging from 10% to 90% (CA 10-90) and NOx emissions first decrease and then increase; and torque, peak in-cylinder pressure (Pmax), and the λ limit first increase and then decrease. The larger the CDIr is, the less influence λ has on the MBT. When CDIr = 15%, the CoVIMEP can be effectively reduced, the engine can still work stably in all lean-burn conditions, and the λ limit will reach the maximum value of 1.73, 19.31% higher than that of the original engine (CDIr = 0). When λ = 1.1, CO emissions decrease the most and HC emissions decrease the least. At this time, CO and HC emissions decrease by 1.56 vol % and 30 ppm, respectively, on average for every 0.1 decrease in λ. For CA 10-90, torque, and Pmax, λ = 1.1, 15% CDI, and 85% EPI is the optimal combination under lean-burn conditions. When CDIr ≥ 15%, NOx emissions are at an ideal level. Under lean-burn conditions, direct-injection CNG can form a good stratified natural gas/ethanol mixture in the cylinder, effectively improving the engine's power and stability and reducing emissions. The λ = 1.1, 15% CDI, 85% EPI combination provides a cutting-edge and outstanding solution for a natural gas/ethanol combined injection SI engine.

Transcriptomic Characterization of Miscanthus sacchariflorus × M. lutarioriparius and Its Implications for Energy Crop Development in the Semiarid Mine Area.

Miscanthus interspecific hybrids have been proved to have better adaptability in marginal lands than their parents. Miscanthus sacchariflorus and Miscanthus lutarioriparius were used as the parents to develop hybrids. We performed the transcriptome for 110 F1 hybrids of Miscanthus sacchariflorus × Miscanthus lutarioriparius and their parents that had been established on the Loess Plateau mine area, to estimate the population's genetic expression variation, and illuminate the adaptive mechanism of the F1 population. The result speculated that the F1 population has mainly inherited the stress response metabolic pathway of its female parent (M. sacchariflorus), which may be responsible for its higher environmental adaptability and biomass yield compared with male parents. Based on PopART, we assembled a leaf reference transcriptome for M. sacchariflorus (LRTMS) and obtained 8116 high-quality transcripts. When we analyze the differential expression of genes between F1 population and parent, 39 and 56 differentially expressed genes were screened out in the female parent and male parent, respectively. The enrichment analysis showed that pathways of carbohydrate metabolism, lipid metabolism, biosynthesis of secondary metabolites and circadian rhythm-plant played a key role in resisting the harsh environment. The carbohydrate metabolism and lipid metabolism were also significantly enriched, and the synthesis of these substances facilitated the yield. The results provided an insight into breeding Miscanthus hybrids more suited to the harsh environment of the Loess Plateau.

Development of energy plants from hybrids between Miscanthus sacchariflorus and M. lutarioriparius grown on reclaimed mine land in the Loess Plateau of China.

Miscanthus, a promising bioenergy plant, has a high biomass yield with high cellulose content suitable for biofuel production. However, harsh climatic and poor soil conditions, such as barren lands or abandoned mines, pose a challenge to the survival and yield of Miscanthus feedstock on the marginal land. The selection from the interspecific hybrids of Miscanthus might combine high survival rates and high yield, which benefits energy crop development in multi-stressful environments. A total of 113 F1 hybrids between Miscanthus sacchariflorus and M. lutarioriparius together with the parents were planted and evaluated for multiple morphological and physiological traits on the mine land of the Loess Plateau of China. The majority of hybrids had higher establishment rates than M. sacchariflorus while M. lutarioriparius failed to survive for the first winter. Nearly all hybrid genotypes outperformed M. lutarioriparius for yield-related traits including plant height, tiller number, tiller diameter, and leaf area. The average biomass of the hybrids was 20 times higher than that of surviving parent, M. sacchariflorus. Furthermore, the photosynthetic rates and water use efficiency of the hybrids were both significantly higher than those of the parents, which might be partly responsible for their higher yield. A total of 29 hybrids with outstanding traits related to yield and stress tolerance were identified as candidates. The study investigated for the first time the hybrids between local individuals of M. sacchariflorus and high-biomass M. lutarioriparius, suggesting that this could be an effective approach for high-yield energy crop development on vast of marginal lands.

Water Use Efficiency and Stress Tolerance of the Potential Energy Crop Miscanthus lutarioriparius Grown on the Loess Plateau of China.

As a potential energy crop with high biomass yield, Miscanthus lutarioriparius (M. lutarioriparius), endemic to the Long River Range in central China, needs to be investigated for its acclimation to stressful climatic and soil conditions often found on the marginal land. In this study, traits related to acclimation and yield, including survival rates, plant height (PH), stem diameter (SD), tiller number (TN), water use efficiency (WUE), and photosynthetic rates (A), were examined for 41 M. lutarioriparius populations that transplanted to the arid and cold Loess Plateau of China. The results showed that the average survival rate of M. lutarioriparius populations was only 4.16% over the first winter but the overwinter rate increased to 35.03% after the second winter, suggesting that plants having survived the first winter could have acclaimed to the low temperature. The strikingly high survival rates over the second winter were found to be 95.83% and 80.85%, respectively, for HG18 and HG39 populations. These populations might be especially valuable for the selection of energy crops for such an area. Those individuals surviving for the two consecutive winters showed significantly higher WUE than those measured after the first winter. The high WUE and low stomatal conductance (gs) observed in survived individuals could have been responsible for their acclimation to this new and harsh environment. A total of 61 individuals with productive growth traits and strong resistance to cold and drought were identified for further energy crop development. This study showed that the variation of M. lutarioriparius held great potential for developing energy crops following continuous field selection.

Chromosome-scale assembly and analysis of biomass crop Miscanthus lutarioriparius genome.

Miscanthus, a rhizomatous perennial plant, has great potential for bioenergy production for its high biomass and stress tolerance. We report a chromosome-scale assembly of Miscanthus lutarioriparius genome by combining Oxford Nanopore sequencing and Hi-C technologies. The 2.07-Gb assembly covers 96.64% of the genome, with contig N50 of 1.71 Mb. The centromere and telomere sequences are assembled for all 19 chromosomes and chromosome 10, respectively. Allotetraploid origin of the M. lutarioriparius is confirmed using centromeric satellite repeats. The tetraploid genome structure and several chromosomal rearrangements relative to sorghum are clearly demonstrated. Tandem duplicate genes of M. lutarioriparius are functional enriched not only in terms related to stress response, but cell wall biosynthesis. Gene families related to disease resistance, cell wall biosynthesis and metal ion transport are greatly expanded and evolved. The expansion of these families may be an important genomic basis for the enhancement of remarkable traits of M. lutarioriparius.

Increased expression diversity buffers the loss of adaptive potential caused by reduction of genetic diversity in new unfavourable environments.

Mechanisms underlying adaptation to rapid environmental change are issues in evolutionary biology. It is widely accepted that reduction in genetic diversity when suddenly exposed to an unfavourable environment limits the adaptive potential of populations. With growing empirical evidence that expression diversity is likely to increase in the new environment, the role that expression diversity plays in adaptation needs to be theorized. Here, we first established a negative exponential relationship between expression diversity and genetic diversity using a phenomenological differential equation. We then derived a complex trade-off relationship between the changes of expression and genetic diversity, which followed a combination of exponential functions. Furthermore, we found the increase in expression diversity could buffer the loss of adaptive potential as genetic diversity decreased to a certain extent. These theoretical deductions were validated by transcriptomic data of Miscanthus lutarioriparius grown in two experimental fields and supported by good fit and random simulation. These results suggest that increased expression diversity may compensate the loss of genetic diversity and allow the populations to maintain a certain level of phenotypic variation to cope with sudden environmental change. This may buffer the quick diminishing of adaptive potential and consequently increases the change of adaptation to the new environment.

N2O and CH4 emission from Miscanthus energy crop fields in the infertile Loess Plateau of China.

BACKGROUND: The greenhouse gas (GHG) mitigation is one of the most important environmental benefits of using bioenergy replacing fossil fuels. Nitrous oxide (N2O) and methane (CH4) are important GHGs and have drawn extra attention for their roles in global warming. Although there have been many works of soil emissions of N2O and CH4 from bioenergy crops in the field scale, GHG emissions in large area of marginal lands are rather sparse and how soil temperature and moisture affect the emission potential remains unknown. Therefore, we sought to estimate the regional GHG emission based on N2O and CH4 releases from the energy crop fields. RESULTS: Here we sampled the top soils from two Miscanthus fields and incubated them using a short-term laboratory microcosm approach under different conditions of typical soil temperatures and moistures. Based on the emission measurements of N2O and CH4, we developed a model to estimate annual regional GHG emission of Miscanthus production in the infertile Loess Plateau of China. The results showed that the N2O emission potential was 0.27 kg N ha-1 year-1 and clearly lower than that of croplands and grasslands. The CH4 uptake potential was 1.06 kg C ha-1 year-1 and was slightly higher than that of croplands. Integrated with our previous study on the emission of CO2, the net greenhouse effect of three major GHGs (N2O, CH4 and CO2) from Miscanthus fields was 4.08 t CO2eq ha-1 year-1 in the Loess Plateau, which was lower than that of croplands, grasslands and shrub lands. CONCLUSIONS: Our study revealed that Miscanthus production may hold a great potential for GHG mitigation in the vast infertile land in the Loess Plateau of China and could contribute to the sustainable energy utilization and have positive environmental impact on the region.

Coexpression network revealing the plasticity and robustness of population transcriptome during the initial stage of domesticating energy crop Miscanthus lutarioriparius.

KEY MESSAGE: Coexpression network revealing genes with Co-variation Expression pattern (CE) and those with Top rank of Expression fold change (TE) played different roles in responding to new environment of Miscanthus lutarioriparius. Variation in gene expression level, the product of genetic and/or environmental perturbation, determines the robustness-to-plasticity spectrum of a phenotype in plants. Understanding how expression variation of plant population response to a new field is crucial to domesticate energy crops. Weighted Gene Coexpression Network Analysis (WGCNA) was used to explore the patterns of expression variation based on 72 Miscanthus lutarioriparius transcriptomes from two contrasting environments, one near the native habitat and the other in one harsh domesticating region. The 932 genes with Co-variation Expression pattern (CE) and other 932 genes with Top rank of Expression fold change (TE) were identified and the former were strongly associated with the water use efficiency (r ≥ 0.55, P ≤ 10-7). Functional enrichment of CE genes were related to three organelles, which well matched the annotation of twelve motifs identified from their conserved noncoding sequence; while TE genes were mostly related to biotic and/or abiotic stress. The expression robustness of CE genes with high genetic diversity kept relatively stable between environments while the harsh environment reduced the expression robustness of TE genes with low genetic diversity. The expression plasticity of CE genes was increased less than that of TE genes. These results suggested that expression variation of CE genes and TE genes could account for the robustness and plasticity of acclimation ability of Miscanthus, respectively. The patterns of expression variation revealed by transcriptomic network would shed new light on breeding and domestication of energy crops.

Population transcriptomic characterization of the genetic and expression variation of a candidate progenitor of Miscanthus energy crops.

The use of transcriptome data in the study of the population genetics of a species can capture faint signals of both genetic variation and expression variation and can provide a broad picture of a species' genomic response to environmental conditions. In this study, we characterized the genetic and expression diversity of Miscanthus lutarioriparius by comparing more than 16,225 transcripts obtained from 78 individuals, belonging to 10 populations distributed across the species' entire geographic range. We only observed a low level of nucleotide diversity (π = 0.000434) among the transcriptome data of these populations, which is consistent with highly conserved sequences of functional elements and protein-coding genes captured with this method. Tests of population divergence using the transcriptome data were consistent with previous microsatellite data but proved to be more sensitive, particularly if gene expression variation was considered as well. For example, the analysis of expression data showed that genes involved in photosynthetic processes and responses to temperature or reactive oxygen species stimuli were significantly enriched in certain populations. This differential gene expression was primarily observed among populations and not within populations. Interestingly, nucleotide diversity was significantly negatively correlated with expression diversity within populations, while this correlation was positive among populations. This suggests that genetic and expression variation play separate roles in adaptation and population persistence. Combining analyses of genetic and gene expression variation represents a promising approach for studying the population genetics of wild species and may uncover both adaptive and nonadaptive processes.

Haplotypes Phased from Population Transcriptomes Detecting Selection in the Initial Adaptation of Miscanthus lutarioriparius to Stressful Environments.

Adaptation is a characteristic that enhances the survival or reproduction of organisms; selection is the critical process leading to adaptive evolution. Therefore, detecting selection is important in studying evolutionary biology. Changes in allele frequency are fundamental to adaptive evolution. The allele frequency of entire genes at the genomic scale is more intensive and precise for analyzing selection effects, compared with simple sequence repeat and single nucleotide polymorphism (SNP) alleles from nuclear gene fragments. Here, we analyzed 29,094 SNPs derived from 80 individuals of 14 L. Liou ex S.L. Chen & Renvoize populations planted near their native habitat (Jiangxia, Hubei Province, JH) and a stressful environment (Qingyang, Gansu Province, QG) to detect selection during initial adaptation. The nucleotide diversity of over 60% of genes was decreased in QG compared with JH, suggesting that most genes were undergoing selection in the stressful environment. We explored a new approach based on haplotype data inferred from RNA-seq data to analyze the change in frequency between two sites and to detect selection signals. In total, 402 and 51 genes were found to be targets of positive and negative selection, respectively. Among these candidate genes, the enrichment of abiotic stress-response genes and photosynthesis-related genes might have been responsible for establishment in the stressful environment. This is the first study assessing the change in allele frequency at the genomic level during adaptation. The method in which allele frequency detects selection during initial adaptation using population RNA-seq data would be useful for developing evolutionary biology.

Systematic comparison of lncRNAs with protein coding mRNAs in population expression and their response to environmental change.

BACKGROUND: Long non-coding RNA (lncRNA) is a class of non-coding RNA with important regulatory roles in biological process of organisms. The systematic comparison of lncRNAs with protein coding mRNAs in population expression and their response to environmental change are still poorly understood. Here we identified 17,610 lncRNAs and calculated their expression levels based on RNA-seq of 80 individuals of Miscanthus lutarioriparius from two environments, the nearly native habitats and transplanted field, respectively. RESULTS: LncRNAs had significantly higher expression diversity and lower expression frequency in population than protein coding mRNAs in both environments, which suggested that lncRNAs may experience more relaxed selection or divergent evolution in population compared with protein coding RNAs. In addition, the increase of expression diversity for lncRNAs was always significantly higher and the magnitude of fold change of expression in new stress environment was significantly larger than protein-coding mRNAs. These results suggested that lncRNAs may be more sensitive to environmental change than protein-coding mRNAs. Analysis of environment-robust and environment-specific lncRNA-mRNA co-expression network between two environments revealed the characterization of lncRNAs in response to environmental change. Furthermore, candidate lncRNAs contributing to water use efficiency (WUE) identified based on the WUE-lncRNA-mRNA co-expression network suggested the roles of lncRNAs in response to environmental change. CONCLUSION: Our study provided a comprehensive understanding of expression characterization of lncRNAs in population for M. lutarioriparius under field condition, which would be useful to explore the roles of lncRNAs and could accelerate the process of adaptation in new environment for many plants.

Sustainable bioenergy production with little carbon debt in the Loess Plateau of China.

BACKGROUND: As a key strategy for mitigating global climate change, bioenergy production by reducing CO2 emissions plays an important role in ensuring sustainable development. However, land-use change by converting natural ecosystems into energy crop field could create a carbon debt at the beginning. Thus, the potential carbon debt calculation is necessary for determining a promising bioenergy crop production, especially in the region rich of marginal land. RESULTS: Here, we used high-resolution historical land-use data to identify the marginal land available and to evaluate the carbon debt of planting Miscanthus in the Loess Plateau, China. We found that there were 27.6 Mha for energy production and 9.7 Mha for ecological restoration, with total annual production of 0.41 billion tons of biomass. We also found that soil carbon sequestration and total CO2 mitigation were 9.3 Mt C year(-1) and 542 Mt year(-1), respectively. More importantly, the result showed that planting Miscanthus on marginal land in the Loess Plateau only took 0.97 years on average to repay the carbon debt. CONCLUSIONS: Our study demonstrated that Miscanthus production in suitable marginal land in the Loess Plateau can offer considerable renewable energy and mitigate climate change with little carbon debt. These results suggested that bioenergy production in the similar arid and semiarid region worldwide would contribute to carbon sequestration in the context of rapid climate change.

Population transcriptomics uncovers the regulation of gene expression variation in adaptation to changing environment.

Expression variation plays an important role in plant adaptation, but little is known about the factors impacting the expression variation when population adapts to changing environment. We used RNA-seq data from 80 individuals in 14 Miscanthus lutarioriparius populations, which were transplanted into a harsh environment from native habitat, to investigate the expression level, expression diversity and genetic diversity for genes expressed in both environments. The expression level of genes with lower expression level or without SNP tended to be more changeable in new environment, which suggested highly expressed genes experienced stronger purifying selection than those at lower level. Low proportion of genes with population effect confirmed the weak population structure and frequent gene flow in these populations. Meanwhile, the number of genes with environment effect was the most frequent compared with that with population effect. Our results showed that environment and genetic diversity were the main factors determining gene expression variation in population. This study could facilitate understanding the mechanisms of global gene expression variation when plant population adapts to changing environment.

The Coordination of Gene Expression within Photosynthesis Pathway for Acclimation of C4 Energy Crop Miscanthus lutarioriparius.

As a promising candidate for the second-generation C4 energy crop, Miscanthus lutarioriparius has well acclimated to the water-limited and high-light Loess Plateau in China by improving photosynthesis rate and water use efficiency (WUE) compared to its native habitat along Yangtze River. Photosynthetic genes were demonstrated as one major category of the candidate genes underlying the physiological superiority. To further study how photosynthetic genes interact to improve the acclimation potential of M. lutarioriparius, population expression patterns within photosynthesis pathway were explored between one mild environment and one harsh environment. We found that 108 transcripts in assembled transcriptome of M. lutarioriparius were highly similar to genes in three Kyoto Encyclopedia of Genes and Genomes (KEGG) photosynthesis pathways of sorghum and maize. Phylogenetic analyses using sorghum, maize, rice, and Arabidopsis genes of dark reaction identified 23 orthologs and 30 paralogs of M. lutarioriparius photosynthetic genes. These genes were also clustered into two kinds of expression pattern. 87% of transcripts in dark reaction were up-regulated and all 14 chloroplast-encoded transcripts in light reaction increased degradation in the harsh environment compared to the mild environment. Moreover, 80.8% of photosynthetic transcripts were coordinated at transcription level under the two environments. Interestingly, LHCI and PSI were significantly correlated with F-ATPase and C4 cycle. Overall, this study indicates the coordinated expression between cyclic electron transport (consisting of LHCI, PSI, and ATPase) and CO2-concentrating mechanism (C4 cycle) could account for photosynthesis plasticity on M. lutarioriparius acclimation potential.

Multiple origins of BBCC allopolyploid species in the rice genus (Oryza).

In the rice genus (Oryza), about one half of the species are allopolyploids. These species are not only important resources for rice breeding but also provide a unique opportunity for studying evolution of polyploid species. In the present study, we sequenced four biparentally inherited nuclear loci and three maternally inherited chloroplast fragments from all diploid and tetraploid species with the B- and C-genome types in this genus. We detected at least three independent origins of three BC-genome tetraploid species. Specifically, the diploid O. punctata (B-genome) and O. officinalis (C-genome) were the parental progenitors of O. minuta and O. malampuzhaensis with O. punctata being the maternal donors, whereas the diploid O. punctata and O. eichingeri (C-genome) were the progenitors of tetraploid O. punctata with O. punctata being the paternal donor. Our relaxed clock analyses suggest that all the BBCC species originated within the last one million years, which is coincident with the severe climate oscillations occurred during the last ice age, implying the potential impact of climate change on their formations and dispersals. In addition, our results support previous taxonomic arguments that the tetraploid O. punctata might be better treated as a separate species (O. schweinfurthiana).

An-2 Encodes a Cytokinin Synthesis Enzyme that Regulates Awn Length and Grain Production in Rice.

A wide range of morphological and physiological traits have changed between cultivated rice Oryza sativa and wild rice Oryza rufipogon under domestication. Here, we report cloning of the An-2 gene, encoding the Lonely Guy Like protein 6 (OsLOGL6), which catalyzes the final step of cytokinin synthesis in O. rufipogon. The near-isogenic line harboring a wild allele of An-2 in the genetic background of the awnless indica Guangluai 4 shows that An-2 promotes awn elongation by enhancing cell division, but decreases grain production by reducing grains per panicle and tillers per plant. We reveal that a genetic variation in the An-2 locus has a large impact on reducing awn length and increasing tiller and grain numbers in domesticated rice. Analysis of gene expression patterns suggests that An-1 regulates the formation of awn primordial, and An-2 promotes awn elongation. Nucleotide diversity of the An-2 locus in cultivated rice was found to be significantly reduced compared with that of wild rice, suggesting that the An-2 locus was subjected to artificial selection. We therefore propose that the selection of genetic variation in An-2 was due to reduced awn length and increased grain yield in cultivated rice.

Transcriptome-wide characterization of candidate genes for improving the water use efficiency of energy crops grown on semiarid land.

Understanding the genetic basis of water use efficiency (WUE) and its roles in plant adaptation to a drought environment is essential for the production of second-generation energy crops in water-deficit marginal land. In this study, RNA-Seq and WUE measurements were performed for 78 individuals of Miscanthus lutarioriparius grown in two common gardens, one located in warm and wet Central China near the native habitats of the species and the other located in the semiarid Loess Plateau, the domestication site of the energy crop. The field measurements showed that WUE of M. lutarioriparius in the semiarid location was significantly higher than that in the wet location. A matrix correlation analysis was conducted between gene expression levels and WUE to identify candidate genes involved in the improvement of WUE from the native to the domestication site. A total of 48 candidate genes were identified and assigned to functional categories, including photosynthesis, stomatal regulation, protein metabolism, and abiotic stress responses. Of these genes, nearly 73% were up-regulated in the semiarid site. It was also found that the relatively high expression variation of the WUE-related genes was affected to a larger extent by environment than by genetic variation. The study demonstrates that transcriptome-wide correlation between physiological phenotypes and expression levels offers an effective means for identifying candidate genes involved in the adaptation to environmental changes.

Population transcriptomics reveals a potentially positive role of expression diversity in adaptation.

While it is widely accepted that genetic diversity determines the potential of adaptation, the role that gene expression variation plays in adaptation remains poorly known. Here we show that gene expression diversity could have played a positive role in the adaptation of Miscanthus lutarioriparius. RNA-seq was conducted for 80 individuals of the species, with half planted in the energy crop domestication site and the other half planted in the control site near native habitats. A leaf reference transcriptome consisting of 18,503 high-quality transcripts was obtained using a pipeline developed for de novo assembling with population RNA-seq data. The population structure and genetic diversity of M. lutarioriparius were estimated based on 30,609 genic single nucleotide polymorphisms. Population expression (Ep ) and expression diversity (Ed ) were defined to measure the average level and the magnitude of variation of a gene expression in the population, respectively. It was found that expression diversity increased while genetic diversity decreased after the species was transplanted from the native habitats to the harsh domestication site, especially for genes involved in abiotic stress resistance, histone methylation, and biomass synthesis under water limitation. The increased expression diversity could have enriched phenotypic variation directly subject to selections in the new environment.

Hollow fiber-based Fabry-Perot cavity for liquid surface tension measurement.

A Fabry-Perot (FP) cavity, formed by a single-mode fiber (SMF) fusion spliced with a section of hollow fiber, was proposed and experimentally used as a sensor for liquid surface tension measurement. Due to the FP cavity length changing with different liquids, the surface tension can be measured by monitoring the interference wavelength spacing. In our experiment, five kinds of liquids were measured, and the relationship between wavelength spacing and liquid surface tension was obtained. The experimental results show that the achieved sensitivity to liquid surface tension is 30.96 nm/(N/m), and the maximum error is 5.43%.

Multiple species of wild tree peonies gave rise to the 'king of flowers', Paeonia suffruticosa Andrews.

The origin of cultivated tree peonies, known as the 'king of flowers' in China for more than 1000 years, has attracted considerable interest, but remained unsolved. Here, we conducted phylogenetic analyses of explicitly sampled traditional cultivars of tree peonies and all wild species from the shrubby section Moutan of the genus Paeonia based on sequences of 14 fast-evolved chloroplast regions and 25 presumably single-copy nuclear markers identified from RNA-seq data. The phylogeny of the wild species inferred from the nuclear markers was fully resolved and largely congruent with morphology and classification. The incongruence between the nuclear and chloroplast trees suggested that there had been gene flow between the wild species. The comparison of nuclear and chloroplast phylogenies including cultivars showed that the cultivated tree peonies originated from homoploid hybridization among five wild species. Since the origin, thousands of cultivated varieties have spread worldwide, whereas four parental species are currently endangered or on the verge of extinction. The documentation of extensive homoploid hybridization involved in tree peony domestication provides new insights into the mechanisms underlying the origins of garden ornamentals and the way of preserving natural genetic resources through domestication.