Genome-Wide Characterization and Functional Validation of the ACS Gene Family in the Chestnut Reveals Its Regulatory Role in Ovule Development.

Ovule abortion significantly contributes to a reduction in chestnut yield. Therefore, an examination of the mechanisms underlying ovule abortion is crucial for increasing chestnut yield. In our previous study, we conducted a comprehensive multiomic analysis of fertile and abortive ovules and found that ACS genes in chestnuts (CmACS) play a crucial role in ovule development. Therefore, to further study the function of ACS genes, a total of seven CmACS members were identified, their gene structures, conserved structural domains, evolutionary trees, chromosomal localization, and promoter cis-acting elements were analyzed, and their subcellular localization was predicted and verified. The spatiotemporal specificity of the expression of the seven CmACS genes was confirmed via qRT-PCR analysis. Notably, CmACS7 was exclusively expressed in the floral organs, and its expression peaked during fertilization and decreased after fertilization. The ACC levels remained consistently greater in fertile ovules than in abortive ovules. The ACSase activity of CmACS7 was identified using the genetic transformation of chestnut healing tissue. Micro Solanum lycopersicum plants overexpressing CmACS7 had a significantly greater rate of seed failure than did wild-type plants. Our results suggest that ovule fertilization activates CmACS7 and increases ACC levels, whereas an overexpression of CmACS7 leads to an increase in ACC content in the ovule prior to fertilization, which can lead to abortion. In conclusion, the present study demonstrated that chestnut ovule abortion is caused by poor fertilization and not by nutritional competition. Optimization of the pollination and fertilization of female flowers is essential for increasing chestnut yield and reducing ovule abortion.

Distribution, causal agents, and infection dynamic of emerging ink disease of sweet chestnut in Southern Switzerland.

Emerging diseases caused by both native and exotic pathogens represent a main threat to forest ecosystems worldwide. The two invasive soilborne pathogens Phytophthora cinnamomi and Phytophthora × cambivora are the causal agents of ink disease, which has been threatening Castanea sativa in Europe for several centuries and seems to be re-emerging in recent years. Here, we investigated the distribution, causal agents, and infection dynamics of ink disease in southern Switzerland. A total of 25 outbreaks were identified, 19 with only P. cinnamomi, 5 with only P. × cambivora, and 1 with both species. Dendrochronological analyses showed that the disease emerged in the last 20-30 years. Infected trees either died rapidly within 5-15 years post-infection or showed a prolonged state of general decline until death. Based on a generalized linear model, the local risk of occurrence of ink disease was increased by an S-SE aspect of the chestnut stand, the presence of a pure chestnut stand, management activities, the proximity of roads and buildings, and increasing annual mean temperature and precipitation. The genetic structure of the local P. cinnamomi population suggests independent introductions and local spread of the pathogen.

Use of retrotransposon based iPBS markers for determination of genetic relationship among some Chestnut Cultivars (Castanea sativa Mill.) in Türkiye.

BACKGROUND: The aim of this study was to reveal the genetic relationships among some economically important chestnut cultivars for Türkiye by using retrotransposon-based inter primer binding site (iPBS) markers. METHODS AND RESULTS: In this study, a total of 19 iPBS markers were used to determine the genetic relationships among 11 chestnut cultivars (Castanea sativa Mill.). In the study, chestnut cultivars named Haciömer, Osmanoglu, Sariaslama, Erfelek, Kemer, Isiklar, Sekerci, Siyah Bursa, Tülü, Bouche De Betizac and Marigoule were the preferred cultivars utilised. Using the online marker efficiency calculator (iMEC), some indices of polymorphism, such as the mean heterozygosity, polymorphism information content, marker index and discriminating power, were determined. In addition, the size ranges of alleles, number of average alleles, number of total alleles, number of polymorphic alleles, and polymorphism rate were determined at a successful level. The chestnut cultivars of Haciömer and Sekerci were determined to be the most similar cultivars with a similarity coefficient value of 0.924, and they formed a subgroup together with the chestnut cultivars Osmanoglu and Erfelek, showing close similarity with these two cultivars. CONCLUSIONS: The use of iPBS markers in chestnuts in Türkiye was carried out for the first time in this study. The power of iPBS markers to evaluate the genetic relationship for our preferred chestnut cultivars was revealed. For this reason, it has emerged that it will be useful in the molecular characterization of both genotypes in natural chestnut populations and chestnut breeding materials such as varieties and cultivars in chestnut breeding programs.

Validation of an Alternative Small Stem Assay for Blight Resistance in Chestnut Seedlings and Recommendations for Broader Use.

We evaluated an alternative small stem assay (AltSSA) for blight resistance in backcross hybrid chestnut trees (Castanea dentata/mollissima). Whereas standard small stem assays (SSAs) are done by inoculating small incisions in stems, in our AltSSA, 4- to 5-mm stems are cut off, and the exposed (living) stem tips are inoculated with discs of Cryphonectria parasitica inoculum and temporarily covered with plastic sleeves. Intended primarily for forward selection, this method was designed to be easy to implement, to consistently induce cankering, and to better enable seedling recovery via the development of lateral shoots from the lower stem. After 90+ days, cankers are evaluated and removed, and seedlings are prepared for out-planting. Previous results showed that AltSSAs performed at least as well as a common SSA method in distinguishing resistant and susceptible types. In this follow-up analysis of 35 lines of backcross seedlings studied in 2020 and 2021, we showed that mean orange zone canker length (OZCL) and a multifactor principal components analysis-based blight resistance index gave results consistent with predictions derived from two methods of blight resistance phenotyping and percentage of American chestnut ancestry of the parents of each line. As expected, based upon the apparent polygenic inheritance of blight resistance in backcross chestnut trees, mean OZCL of backcross families ranged from intermediate (F1 hybrid-level) to low (wild-type American chestnut-level). Consistent with prior results, canker production was near 100%, survivorship after out-planting was very high, and postinoculation stem dieback was not apparently related to the stem tip inoculations. Altogether, these results suggest that the AltSSA is a viable method for early detection of relative blight resistance in seedlings and may enable a reduction in the numbers of trees out-planted and placed under care for long-term evaluation and breeding. Thus, the AltSSA can prevent time, resources, and orchard space from being used on susceptible trees.

Frozen in time: Rangewide genomic diversity, structure, and demographic history of relict American chestnut populations.

American chestnut (Castanea dentata) was once the most economically and ecologically important hardwood species in the eastern United States. In the first half of the 20th century, an exotic fungal pathogen-Cryphonectria parasitica-decimated the species, killing billions of chestnut trees. Two approaches to developing blight-resistant American chestnut populations show promise, but both will require introduction of adaptive genomic diversity from wild germplasm to produce diverse, locally adapted restoration populations. Here we characterize population structure, demographic history, and genomic diversity in a range-wide sample of 384 wild American chestnuts to inform conservation and breeding with blight-resistant varieties. Population structure analyses suggest that the chestnut range can be roughly divided into northeast, central, and southwest populations. Within-population genomic diversity estimates revealed a clinal pattern with the highest diversity in the southwest, which likely reflects bottleneck events associated with Quaternary glaciation. Finally, we identified genomic regions under positive selection within each population, which suggests that defence against fungal pathogens is a common target of selection across all populations. Taken together, these results show that American chestnut underwent a postglacial expansion from the southern portion of its range leading to three extant genetic populations. These populations will serve as management units for breeding adaptive genetic variation into the blight-resistant tree populations for targeted reintroduction efforts.

Transcriptome Analysis Reveals the Regulatory Networks of Cytokinin in Promoting Floral Feminization in Castanea henryi.

Castanea henryi is a monoecious plant with a low female-to-male ratio, which limits its yield. The phytohormone cytokinin (CK) plays a crucial role in flower development, especially gynoecium development. Here, the feminizing effect of CK on the development of C. henryi was confirmed by the exogenous spraying of N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU). Spraying CPPU at 125 mg·L-1 thrice changed the male catkin into a pure female catkin, whereas at 5 mg·L-1 and 25 mg·L-1, only a part of the male catkin was transformed into a female catkin. A comparative transcriptome analysis of male catkins subjected to CPPU was performed to study the mechanism of the role of CKs in sex differentiation. Using Pearson's correlation analysis between hormone content and hormone synthesis gene expression, four key genes, LOG1, LOG3, LOG7 and KO, were identified in the CK and GA synthesis pathways. Moreover, a hub gene in the crosstalk between JA and the other hormone signaling pathways, MYC2, was identified, and 15 flowering-related genes were significantly differentially expressed after CPPU treatment. These results suggest that CK interacts with other phytohormones to determine the sex of C. henryi, and CK may directly target floral organ recognition genes to control flower sex.

Evolution of Castanea in North America: restriction-site-associated DNA sequencing and ecological modeling reveal a history of radiation, range shifts, and disease.

PREMISE: Although chestnuts and chinquapins are some of the best known and most widely loved of any plants in North America, relatively little genomic sequencing has been done, and much is still unknown about their evolution. METHODS: We used double-digest restriction-site-associated DNA (ddRAD) sequencing data to infer the species-level phylogeny for Castanea and assess the phylogeography of the North American species using samples collected from populations that span the full extent of the species' ranges. We also constructed species distribution models using digitized herbarium specimens and observational data from field surveys. RESULTS: We identified strong population structure within Castanea dentata (American chestnut) that reflects a stepwise northern migration since the last glacial maximum. Our species distribution models further confirmed this scenario and matched closely with the Castanea fossil pollen record. We also found significant structure within the Castanea pumila lineage, most notably a genetic cluster that corresponds to the frequently recognized Castanea pumila var. ozarkensis. CONCLUSIONS: The two North American Castanea species have contrasting patterns of population structure, but each is typical of plant phylogeography in North America. Within the C. pumila complex, we found novel genetic structure that provides new insights about C. pumila taxonomy. Our results also identified a series of distinctive populations that will be valuable in ongoing efforts to conserve and restore chestnuts and chinquapins in North America.

Adaptive Evolution of Chalcone Isomerase Superfamily in Fagaceae.

Chalcone Isomerase (CHI) catalyzes the biosynthesis of flavonoids and secondary metabolism in plants. Currently, there is no systematic analysis of CHIs gene family in Fagaceae which is available. In this study, twenty-two CHI proteins were identified in five species of the Fagaceae family. The CHI superfamily in Fagaceae can be classified into three subfamilies and five groups using phylogenetic analysis, analysis of physicochemical properties, and structural prediction. Results indicated that serine (Ser) and isoleucine (Ile) residues determine the substrate preferred by active Type I Fagaceae CHI, and the chalcone isomerase-like (CHIL) of Fagaceae had active site residues. Adaptive analysis of CHIs showed that CHIs are subject to selection pressure. The active CHI gene of Fagaceae was located in the cytoplasm, and it had the typical gene structure of CHI and contains four exons. All the twenty-two identified CHIs had the conserved domain motif 3, and the different groups had their own structural characteristics. In the process of fatty acid binding protein (FAP) evolution to CHIL and CHI, the physical and chemical properties of proteins also had significant differences in addition to changes in protein functions.

Transcriptome characterization and expression profiling in chestnut cultivars resistant or susceptible to the gall wasp Dryocosmus kuriphilus.

The oriental gall wasp Dryocosmus kuriphilus represents a limiting pest for the European Chestnut (Castanea sativa, Fagaceae) as it creates severe yield losses. The European Chestnut is a deciduous tree, having major social, economic and environmental importance in Southern Europe, covering an area of 2.53 million hectares, including 75,000 ha devoted to fruit production. Cultivars show different susceptibility and very few are resistant to gall wasp. To deeply investigate the plant response and understand which factors can lead the plant to develop or not the gall, the study of transcriptome is basic (fundamental). To date, little transcriptomic information are available for C. sativa species. Hence, we present a de novo assembly of the chestnut transcriptome of the resistant Euro-Japanese hybrid 'Bouche de Bétizac' (BB) and the susceptible cultivar 'Madonna' (M), collecting RNA from buds at different stages of budburst. The two transcriptomes were assembled into 34,081 (BB) and 30,605 (M) unigenes, respectively. The former was used as a reference sequence for further characterization analyses, highlighting the presence of 1444 putative resistance gene analogs (RGAs) and about 1135 unigenes, as putative MiRNA targets. A global quantitative transcriptome profiling comparing the resistant and the susceptible cultivars, in the presence or not of the gall wasp, revealed some GO enrichments as "response to stimulus" (GO:0050896), and "developmental processes" (e.g., post-embryonic development, GO:0009791). Many up-regulated genes appeared to be transcription factors (e.g., RAV1, AP2/ERF, WRKY33) or protein regulators (e.g., RAPTOR1B) and storage proteins (e.g., LEA D29) involved in "post-embryonic development". Our analysis was able to provide a large amount of information, including 7k simple sequence repeat (SSR) and 335k single-nucleotide polymorphism (SNP)/INDEL markers, and generated the first reference unigene catalog for the European Chestnut. The transcriptome data for C. sativa will contribute to understand the genetic basis of the resistance to gall wasp and will provide useful information for next molecular genetic studies of this species and its relatives.

Transcriptome Analysis and Identification of Genes Associated with Starch Metabolism in Castanea henryi Seed (Fagaceae).

Starch is the most important form of carbohydrate storage and is the major energy reserve in some seeds, especially Castanea henryi. Seed germination is the beginning of the plant's life cycle, and starch metabolism is important for seed germination. As a complex metabolic pathway, the regulation of starch metabolism in C. henryi is still poorly understood. To explore the mechanism of starch metabolism during the germination of C. henryi, we conducted a comparative gene expression analysis at the transcriptional level using RNA-seq across four different germination stages, and analyzed the changes in the starch and soluble sugar contents. The results showed that the starch content increased in 0-10 days and decreased in 10-35 days, while the soluble sugar content continuously decreased in 0-30 days and increased in 30-35 days. We identified 49 candidate genes that may be associated with starch and sucrose metabolism. Three ADP-glucose pyrophosphorylase (AGPase) genes, two nucleotide pyrophosphatase/phosphodiesterases (NPPS) genes and three starch synthases (SS) genes may be related to starch accumulation. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the expression levels of these genes. Our study combined transcriptome data with physiological and biochemical data, revealing potential candidate genes that affect starch metabolism during seed germination, and provides important data about starch metabolism and seed germination in seed plants.

Approximate Bayesian computation analysis of EST-associated microsatellites indicates that the broadleaved evergreen tree Castanopsis sieboldii survived the Last Glacial Maximum in multiple refugia in Japan.

Climatic changes have played major roles in plants' evolutionary history. Glacial oscillations have been particularly important, but some of their effects on plants' populations are poorly understood, including the numbers and locations of refugia in Asian warm temperate zones. In the present study, we investigated the demographic history of the broadleaved evergreen tree species Castanopsis sieboldii (Fagaceae) during the last glacial period in Japan. We used approximate Bayesian computation (ABC) for model comparison and parameter estimation for the demographic modeling using 27 EST-associated microsatellites. We also performed the species distribution modeling (SDM). The results strongly support a demographic scenario that the Ryukyu Islands and the western parts in the main islands (Kyushu and western Shikoku) were derived from separate refugia and the eastern parts in the main islands and the Japan Sea groups were diverged from the western parts prior to the coldest stage of the Last Glacial Maximum (LGM). Our data indicate that multiple refugia survived at least one in the Ryukyu Islands, and the other three regions of the western and eastern parts and around the Japan Sea of the main islands of Japan during the LGM. The SDM analysis also suggests the potential habitats under LGM climate conditions were mainly located along the Pacific Ocean side of the coastal region. Our ABC-based study helps efforts resolve the demographic history of a dominant species in warm temperate broadleaved forests during and after the last glacial period, which provides a basic model for future phylogeographical studies using this approach.

Chestnuts bred for blight resistance depart nursery with distinct fungal rhizobiomes.

Restoration of the American chestnut (Castanea dentata) is underway using backcross breeding that confers chestnut blight disease resistance from Asian chestnuts (most often Castanea mollissima) to the susceptible host. Successful restoration will depend on blight resistance and performance of hybrid seedlings, which can be impacted by below-ground fungal communities. We compared fungal communities in roots and rhizospheres (rhizobiomes) of nursery-grown, 1-year-old chestnut seedlings from different genetic families of American chestnut, Chinese chestnut, and hybrids from backcross breeding generations as well as those present in the nursery soil. We specifically focused on the ectomycorrhizal (EcM) fungi that may facilitate host performance in the nursery and aid in seedling establishment after outplanting. Seedling rhizobiomes and nursery soil communities were distinct and seedlings recruited heterogeneous communities from shared nursery soil. The rhizobiomes included EcM fungi as well as endophytes, putative pathogens, and likely saprobes, but their relative proportions varied widely within and among the chestnut families. Notably, hybrid seedlings that hosted few EcM fungi hosted a large proportion of potential pathogens and endophytes, with possible consequences in outplanting success. Our data show that chestnut seedlings recruit divergent rhizobiomes and depart nurseries with communities that may facilitate or compromise the seedling performance in the field.

Transcriptome Analysis of Chinese Chestnut (Castanea mollissima Blume) in Response to Dryocosmus kuriphilus Yasumatsu Infestation.

Chinese chestnut (Castanea mollissima Blume) can be infested by Dryocosmus kuriphilus Yasumatsu, resulting in gall formation and yield losses. Research on the control of gall wasps using genomics approaches is rarely reported. We used RNA-seq to investigate the dynamic changes in the genes of a chestnut species (C. mollissima B.) during four gall-formation stages caused by D. kuriphilus. A total of 21,306 genes were annotated by BLAST in databases. Transcriptome comparison between different gall-formation stages revealed many genes that were differentially expressed compared to the control. Among these, 2410, 7373, 6294, and 9412 genes were differentially expressed in four gall-formation stages: initiation stage (A), early growth stage (B), late growth stage (C), and maturation stage (D), respectively. Annotation analysis indicated that many metabolic processes (e.g., phenylpropanoid biosynthesis, secondary metabolism, plant⁻pathogen interaction) were affected. Interesting genes encoding putative components of signal transduction, stress response, and transcription factors were also differentially regulated. These genes might play important roles in response to D. kuriphilus gall formation. These new data on the mechanism by which D. kuriphilus infests chestnuts could help improve chestnut resistance.

Roles of the GA-mediated SPL Gene Family and miR156 in the Floral Development of Chinese Chestnut (Castanea mollissima).

Chestnut (Castanea mollissima) is a deciduous tree species with major economic and ecological value that is widely used in the study of floral development in woody plants due its monoecious and out-of-proportion characteristics. Squamosa promoter-binding protein-like (SPL) is a plant-specific transcription factor that plays an important role in floral development. In this study, a total of 18 SPL genes were identified in the chestnut genome, of which 10 SPL genes have complementary regions of CmmiR156. An analysis of the phylogenetic tree of the squamosa promoter-binding protein (SBP) domains of the SPL genes of Arabidopsis thaliana, Populus trichocarpa, and C. mollissima divided these SPL genes into eight groups. The evolutionary relationship between poplar and chestnut in the same group was similar. A structural analysis of the protein-coding regions (CDSs) showed that the domains have the main function of SBP domains and that other domains also play an important role in determining gene function. The expression patterns of CmmiR156 and CmSPLs in different floral organs of chestnut were analyzed by real-time quantitative PCR. Some CmSPLs with similar structural patterns showed similar expression patterns, indicating that the gene structures determine the synergy of the gene functions. The application of gibberellin (GA) and its inhibitor (Paclobutrazol, PP333) to chestnut trees revealed that these exert a significant effect on the number and length of the male and female chestnut flowers. GA treatment significantly increased CmmiR156 expression and thus significantly decreased the expression of its target gene, CmSPL6/CmSPL9/CmSPL16, during floral bud development. This finding indicates that GA might indirectly affect the expression of some of the SPL target genes through miR156. In addition, RNA ligase-mediated rapid amplification of the 5' cDNA ends (RLM-RACE) experiments revealed that CmmiR156 cleaves CmSPL9 and CmSPL16 at the 10th and 12th bases of the complementary region. These results laid an important foundation for further study of the biological function of CmSPLs in the floral development of C. mollissima.

Molecular genetic and biochemical evidence for adaptive evolution of leaf abaxial epicuticular wax crystals in the genus Lithocarpus (Fagaceae).

BACKGROUND: Leaf epicuticular wax is an important functional trait for physiological regulation and pathogen defense. This study tests how selective pressure may have forced the trait of leaf abaxial epicuticular wax crystals (LAEWC) and whether the presence/absence of LAEWC is associated with other ecophysiological traits. Scanning Electron Microscopy was conducted to check for LAEWC in different Lithocarpus species. Four wax biosynthesis related genes, including two wax backbone genes ECERIFERUM 1 (CER1) and CER3, one regulatory gene CER7 and one transport gene CER5, were cloned and sequenced. Ecophysiological measurements of secondary metabolites, photosynthesis, water usage efficiency, and nutrition indices were also determined. Evolutionary hypotheses of leaf wax character transition associated with the evolution of those ecophysiological traits as well as species evolution were tested by maximum likelihood. RESULTS: Eight of 14 studied Lithocarpus species have obvious LAEWC appearing with various types of trichomes. Measurements of ecophysiological traits show no direct correlations with the presence/absence of LAEWC. However, the content of phenolic acids is significantly associated with the gene evolution of the wax biosynthetic backbone gene CER1, which was detected to be positively selected when LAEWC was gained during the late-Miocene-to-Pliocene period. CONCLUSIONS: Changes of landmass and vegetation type accelerated the diversification of tropical and subtropical forest trees and certain herbivores during the late Miocene. As phenolic acids were long thought to be associated with defense against herbivories, co-occurrence of LAEWC and phenolic acids may suggest that LAEWC might be an adaptive defensive mechanism in Lithocarpus.

Development of 13 microsatellite markers for Castanopsis tribuloides (Fagaceae) using next-generation sequencing.

Catanopsis tribuloides is a climax tree species commonly distributed in evergreen forests and has been used to restore degraded areas in northern Thailand. To aid in study of genetic diversity of the species, microsatellite markers, which are specific to C. tribuloides, were developed using whole genome sequencing by next-generation sequencing technology. The primers for microsatellite were developed and screened for optimal annealing temperature by PCR assay. The loci primers specific with C. tribuloides, 13 polymorphic microsatellite primers were successfully developed. The results from genetic information analyzing showed the number of alleles presented were between 2 and 24. Accordingly, the expected and observed heterozygosity obtained were between 0.298 and 0.920 and 0.364 to 1.000, respectively. Null allele frequency was presented 0.000-0.199. Genetic information was generated 10 loci primers significantly deviated from Hardy-Weinberg Equilibrium. All 13 primer pairs of loci were not significant with linkage disequilibrium. A set of microsatellite markers in this study could be applied to gene flow, genetic structure and population genetic studies in the future.

Molecular characterization of Fagaceae species using inter-primer binding site (iPBS) markers.

Retrotransposons (RTNs) contribute for genome evolution, influencing its size and structure. We investigated the utility of the RTN-based markers inter-primer binding site (iPBS) for the molecular characterization of 25 Fagaceae species from genera Castanea, Fagus and Quercus. The assessment of genetic diversity, relationships and structure, as well as taxonomic classification of Fagaceae based on molecular data is important for definition of conservation, forestry management strategies and discrimination among natural hybrids and their parents since natural hybridization may increase with the climate changes. Here, iPBS primers designed by other authors were tested alone and combined. Some of them were discriminative, revealed polymorphism within and among taxa allowing the production of a total of 150 iPBS markers. In addition, several monomorphic iPBS markers were also amplified in each taxon. The UPGMA dendrogram based on the pooled iPBS data revealed 27% of genetic similarity among species. The individuals were clustered per genus and most of the oaks per infrageneric group corroborating the adopted taxonomy. Globally, the iPBS markers demonstrated suitability for DNA fingerprinting, determination of phylogenies and taxonomic discrimination in Fagaceae, and could constitute a useful and alternative tool for germplasm characterization, and for definition of conservation strategies and forestry management. Moreover, these markers would be useful for fingerprinting natural hybrids that share morphological similarities with their parents. Since iPBS markers could also enable insights about RTNs evolution, an eventual correlation among iPBS polymorphism, variability of RTN insertions and/or genome size in Fagaceae is discussed.

Patterns of genomic diversification reflect differences in life history and reproductive biology between figs (Ficus) and the stone oaks (Lithocarpus).

One of the remarkable aspects of the tremendous biodiversity found in tropical forests is the wide range of evolutionary strategies that have produced this diversity, indicating many paths to diversification. We compare two diverse groups of trees with profoundly different biologies to discover whether these differences are reflected in their genomes. Ficus (Moraceae), with its complex co-evolutionary relationship with obligate pollinating wasps, produces copious tiny seeds that are widely dispersed. Lithocarpus (Fagaceae), with generalized insect pollination, produces large seeds that are poorly dispersed. We hypothesize that these different reproductive biologies and life history strategies should have a profound impact on the basic properties of genomic divergence within each genus. Using shallow whole genome sequencing for six species of Ficus, seven species of Lithocarpus, and three outgroups, we examined overall genomic diversity, how it is shared among the species within each genus, and the fraction of this shared diversity that agrees with the major phylogenetic pattern. A substantially larger fraction of the genome is shared among species of Lithocarpus, a considerable amount of this shared diversity was incongruent with the general background history of the genomes, and each fig species possessed a substantially larger fraction of unique diversity than Lithocarpus.

[Analysis of transicriptomes and exploring flavonoid biosynthetic pathway genes in Lithocarpus polystachyus].

The sweet taste and health effect of Lithocarpus polystachyus are mainly related flavonoid. To obtain Lithocarpus transcriptome database and flavonoid biosynthesis-related genes, the RNA-Seq techology (Illumina HiSeq 4000) was used to sequence its transcriptome. Six Gb database was assembled after assembly steps, and 41 043 of L. polystachyus unigenes were obtained. With blasting them with 7 data banks, all unigenes were involved in 51 GO-terms and 237 metabolic pathways. And furthermore 28 genes of the flavonoid biosynthesis-related were found. After using the MicroSatallite, 18 161 SSR were obtained, the single-nucleotide-repeated was the richest at 7 346. These data represent abundant messages about transcripts and provide valuable genome data sources in molecular biology of L. polystachyus.

Divergence maintained by climatic selection despite recurrent gene flow: a case study of Castanopsis carlesii (Fagaceae).

Local adaptation to different environments has the potential to maintain divergence between populations despite recurrent gene flow and is an important driver for generating biological diversity. In this study, we investigate the role of adaptation in the maintenance of two parapatric varieties of a forest tree. We used sequence variation of chloroplastic DNA and restriction site-associated DNA to investigate the genetic structure of two varieties of Castanopsis carlesii in subtropical China and relate it to climatic variation. We used niche reconstruction methods to investigate niche differentiation between the two varieties and to estimate the past distribution of this species. A deep divergence was observed between the two varieties, but evidence of introgression and genetic admixture was detected in two phenotypically and geographically intermediate populations. Niche reconstruction suggests that the distribution of the two varieties was disjunct during periods of global cooling and that the two varieties occupy significantly different niches. The genetic structure was mainly driven by environmental factors, and 13 outlier loci under divergent selection were correlated with climatic variation. These results suggest that the two varieties evolved in allopatry and came back into secondary contact after the last glacial maximum and that they are an evolutionary example of divergence maintained by climatic selection despite recurrent gene flow.