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An Illumina Infinium SNP genotyping array was constructed for European white oaks. Six individuals of Quercus petraea and Q. robur were considered for SNP discovery using both previously obtained Sanger sequences across 676 gene regions (1371 in vitro SNPs) and Roche 454 technology sequences from 5112 contigs (6542 putative in silico SNPs). The 7913 SNPs were genotyped across the six parental individuals, full-sib progenies (one within each species and two interspecific crosses between Q. petraea and Q. robur) and three natural populations from south-western France that included two additional interfertile white oak species ( Q. pubescens and Q. pyrenaica). The genotyping success rate in mapping populations was 80.4% overall and 72.4% for polymorphic SNPs. In natural populations, these figures were lower (54.8% and 51.9%, respectively). Illumina genotype clusters with compression (shift of clusters on the normalized x-axis) were detected in ~25% of the successfully genotyped SNPs and may be due to the presence of paralogues. Compressed clusters were significantly more frequent for SNPs showing a priori incorrect Illumina genotypes, suggesting that they should be considered with caution or discarded. Altogether, these results show a high experimental error rate for the Infinium array (between 15% and 20% of SNPs potentially unreliable and 10% when excluding all compressed clusters), and recommendations are proposed when applying this type of high-throughput technique. Finally, results on diversity levels and shared polymorphisms across targeted white oaks and more distant species of the Quercus genus are discussed, and perspectives for future comparative studies are proposed. [ABSTRACT FROM AUTHOR]

We analyzed the genetic mosaic of speciation in two hybridizing Mediterranean white oaks from the Iberian Peninsula (Quercus faginea Lamb. and Quercus pyrenaica Willd.). The two species show ecological divergence in flowering phenology, leaf morphology and composition, and in their basic or acidic soil preferences. Ninety expressed sequence tag-simple sequence repeats (EST-SSRs) and eight nuclear SSRs were genotyped in 96 trees from each species. Genotyping was designed in two steps. First, we used 69 markers evenly distributed over the 12 linkage groups (LGs) of the oak linkage map to confirm the species genetic identity of the sampled genotypes, and searched for differentiation outliers. Then, we genotyped 29 additional markers from the chromosome bins containing the outliers and repeated the multilocus scans. We found one or two additional outliers within four saturated bins, thus confirming that outliers are organized into clusters. Linkage disequilibrium (LD) was extensive; even for loosely linked and for independent markers. Consequently, score tests for association between two-marker haplotypes and the 'species trait' showed a broad genomic divergence, although substantial variation across the genome and within LGs was also observed. We discuss the influence of several confounding effects on neutrality tests and review the evolutionary processes leading to extensive LD. Finally, we examine how LD analyses within regions that contain outlier clusters and quantitative trait loci can help to identify regions of divergence and/or genomic hitchhiking in the light of predictions from ecological speciation theory. [ABSTRACT FROM AUTHOR]

Nucleotide diversity was assessed within nine candidate genes (CGs) (in total 4.6 kb) for the time of bud burst in nine sessile oak (Quercus petraea) populations distributed in central and northern Europe. The sampled populations were selected on the basis of their contrasting times of bud burst observed in common garden experiments (provenance tests). The CGs were selected according to their expression profiles during the transition from quiescent to developing buds and/or their functional role in model plants. The overall nucleotide diversity was large (πtot=6.15 × 10−3; πsilent=11.2 × 10−3), but population differentiation was not larger than for microsatellites. No outlier single-nucleotide polymorphism (SNP) departing from neutral expectation was found among the total of 125 SNPs. These results contrasted markedly with the significant associations that were observed between the CGs and bud burst in segregating populations. Quantitative trait loci (QTLs) for bud burst were identified for 13 year*site seasonal observations in a cloned mapping pedigree. Nineteen QTLs were detected, and QTLs located on linkage groups 2, 5 and 9 contributed repeatedly to more than 12% of the phenotypic variation of the trait. Eight genes were polymorphic in the two parents of the pedigree and could be mapped on the existing genetic map. Five of them located within the confidence intervals of QTLs for bud burst. Interestingly, four of them located within the three QTLs exhibiting the largest contributions to bud burst. [ABSTRACT FROM AUTHOR]

Pedunculate oak and sessile oak are two sympatric interfertile species that exhibit leaf morphological differences. We aimed to detect quantitative trait loci (QTLs) of these traits in order to locate genomic regions involved in species differentiation. A total of 15 leaf morphological traits were assessed in a mixed forest stand composed of Quercus petraea and Q. robur and in a full-sib pedigree of Q. robur. The progeny of the full-sib family were vegetatively propagated in two successive experiments comprising 174 and 216 sibs, and assessments were made on two leaves collected on each of the 1080 and 1530 cuttings corresponding to the two experiments. Traits that exhibited strong species differences in the mixed stand tended also to have higher repeatability values in the mapping population, thus indicating higher genetic control. A genetic map was constructed for QTL detection. Composite interval mapping with the one QTL model was used for QTL detection. From one to three QTLs were detected for 13 traits. In-depth analysis of the QTLs, controlling the five morphological traits that exhibited the highest interspecific differences in the mixed stand, indicated that they were distributed on six linkage groups, with two clusters comprising QTLs of at least two discriminant traits. These results were reinforced when error 1 for QTL detection was set at 5% at the chromosome level, as up to nine clusters could be identified. In conclusion, traits involved in interspecific differentiation of oaks are under polygenic control and widespread in clusters across the genome.Heredity (2004) 92, 20-30, advance online publication, 24 September 2003; doi:10.1038/sj.hdy.6800358 [ABSTRACT FROM AUTHOR]

Abstract The two most common oak species in western Europe, Quercus robur and Quercus petraea, display different ecological behaviours, particularly with respect to their responses to drought. The ecophysiological basis of this niche difference is not understood well. Here we test the hypothesis that these two species present distinct water use efficiencies (WUEs), using the carbon isotope discrimination approach. Leaves and 13 dated ring sequences were sampled in 10 pairs of adult trees growing side by side. Carbon isotope composition was measured on cellulose extracts. In addition, relationships between carbon isotope discrimination and wood anatomy were assessed at the tree level. Quercus robur displayed a 1·0‰ larger isotopic discrimination than Q. petraea, and therefore a lower intrinsic WUE (-13%). This interspecific difference of isotopic discrimination was quite stable with time and independent of tree radial growth and climate fluctuations. A strong positive correlation was observed between average tree values of earlywood vessel surface area and 13C isotopic discrimination. This correlation was even higher with 13C of the 1976 dry year (r = 0·86). These observations led to the hypothesis that hydraulic properties of xylem could exert a constraint on leaf gas exchange, resulting in a larger WUE for individuals with smaller vessel cross-section area. [ABSTRACT FROM AUTHOR]

Abstract This study compares the properties of dominant markers, such as amplified fragment length polymorphisms (AFLPs), with those of codominant multiallelic markers, such as microsatellites, in reconstructing parentage. These two types of markers were used to search for both parents of an individual without prior knowledge of their relationships, by calculating likelihood ratios based on genotypic data, including mistyping. Experimental data on 89 oak trees genotyped for six microsatellite markers and 159 polymorphic AFLP loci were used as a starting point for simulations and tests. Both sets of markers produced high exclusion probabilities, and among dominant markers those with dominant allele frequencies in the range 0.1–0.4 were more informative. Such codominant and dominant markers can be used to construct powerful statistical tests to decide whether a genotyped individual (or two individuals) can be considered as the true parent (or parent pair). Gene flow from outside the study stand (GFO), inferred from parentage analysis with microsatellites, overestimated the true GFO, whereas with AFLPs it was underestimated. As expected, dominant markers are less efficient than codominant markers for achieving this, but can still be used with good confidence, especially when loci are deliberately selected according to their allele frequencies. [ABSTRACT FROM AUTHOR]

The European Beech is the dominant climax tree in most regions of Central Europe and valued for its ecological versatility and hardwood timber. Even though a draft genome has been published recently, higher resolution is required for studying aspects of genome architecture and recombination. Here, we present a chromosome-level assembly of the more than 300 year-old reference individual, Bhaga, from the Kellerwald-Edersee National Park (Germany). Its nuclear genome of 541 Mb was resolved into 12 chromosomes varying in length between 28 and 73 Mb. Multiple nuclear insertions of parts of the chloroplast genome were observed, with one region on chromosome 11 spanning more than 2 Mb which fragments up to 54,784 bp long and covering the whole chloroplast genome were inserted randomly. Unlike in Arabidopsis thaliana , ribosomal cistrons are present in Fagus sylvatica only in four major regions, in line with FISH studies. On most assembled chromosomes, telomeric repeats were found at both ends, while centromeric repeats were found to be scattered throughout the genome apart from their main occurrence per chromosome. The genome-wide distribution of SNPs was evaluated using a second individual from Jamy Nature Reserve (Poland). SNPs, repeat elements and duplicated genes were unevenly distributed in the genomes, with one major anomaly on chromosome 4. The genome presented here adds to the available highly resolved plant genomes and we hope it will serve as a valuable basis for future research on genome architecture and for understanding the past and future of European Beech populations in a changing climate., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Mishra, Ulaszewski, Meger, Aury, Bodénès, Lesur-Kupin, Pfenninger, Da Silva, Gupta, Guichoux, Heer, Lalanne, Labadie, Opgenoorth, Ploch, Le Provost, Salse, Scotti, Wötzel, Plomion, Burczyk and Thines.)

Exotic pathogens cause severe damage in natural populations in the absence of coevolutionary dynamics with their hosts. However, some resistance to such pathogens may occur in naive populations. The objective of this study was to investigate the genetics of this so-called 'exapted' resistance to two pathogens of Asian origin (Erysiphe alphitoides and Phytophthora cinnamomi) in European oak. Host-pathogen compatibility was assessed by recording infection success and pathogen growth in a full-sib family of Quercus robur under controlled and natural conditions. Two high-resolution genetic maps anchored on the reference genome were used to study the genetic architecture of resistance and to identify positional candidate genes. Two genomic regions, each containing six strong and stable quantitative trait loci (QTLs) accounting for 12-19% of the phenotypic variation, were mainly associated with E. alphitoides infection. Candidate genes, especially genes encoding receptor-like-kinases and galactinol synthases, were identified in these regions. The three QTLs associated with P. cinnamomi infection did not colocate with QTLs found for E. alphitoides. These findings provide evidence that exapted resistance to E. alphitoides and P. cinnamomi is present in Q. robur and suggest that the underlying molecular mechanisms involve genes encoding proteins with extracellular signaling functions., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

The tree of life is highly reticulate, with the history of population divergence emerging from populations of gene phylogenies that reflect histories of introgression, lineage sorting and divergence. In this study, we investigate global patterns of oak diversity and test the hypothesis that there are regions of the oak genome that are broadly informative about phylogeny. We utilize fossil data and restriction-site associated DNA sequencing (RAD-seq) for 632 individuals representing nearly 250 Quercus species to infer a time-calibrated phylogeny of the world's oaks. We use a reversible-jump Markov chain Monte Carlo method to reconstruct shifts in lineage diversification rates, accounting for among-clade sampling biases. We then map the > 20 000 RAD-seq loci back to an annotated oak genome and investigate genomic distribution of introgression and phylogenetic support across the phylogeny. Oak lineages have diversified among geographic regions, followed by ecological divergence within regions, in the Americas and Eurasia. Roughly 60% of oak diversity traces back to four clades that experienced increases in net diversification, probably in response to climatic transitions or ecological opportunity. The strong support for the phylogeny contrasts with high genomic heterogeneity in phylogenetic signal and introgression. Oaks are phylogenomic mosaics, and their diversity may in fact depend on the gene flow that shapes the oak genome., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

Oaks are dominant forest tree species widely distributed across the Northern Hemisphere, where they constitute natural resources of economic, ecological, social and historical value. Hybridisation and adaptive introgression have long been thought to be major drivers of their ecological success. Therefore, the maintenance of species barriers remains a key question, given the extent of interspecific gene flow. In this study, we made use of the tremendous genetic variation among four European white oak species (31 million single nucleotide polymorphisms (SNPs)) to infer the evolutionary history of these species, study patterns of genetic differentiation and identify reproductive barriers. We first analysed the ecological and historical relationships among these species and inferred a long-term strict isolation followed by a recent and extensive postglacial contact using approximate Bayesian computation. Assuming this demographic scenario, we then performed backward simulations to generate the expected distributions of differentiation under neutrality to scan their genomes for reproductive barriers. We finally identified important intrinsic and ecological functions driving the reproductive isolation. We discussed the importance of identifying the genetic basis for the ecological preferences between these oak species and its implications for the renewal of European forests under global warming., (© 2019 The Authors New Phytologist © 2019 New Phytologist Trust.)

Reproduction, one of the main components of plant fitness, is highly variable in response to environmental cues, but little is known about the genetic determinism underlying reproduction-related traits in forest tree species. There is therefore an urgent need to characterize the genetic architecture of those traits if we are to predict the evolutionary trajectories of forest populations facing rapidly changing environment and mitigate their impacts. Using a full-sib family of pedunculate oak ( Quercus robur ), we investigated the within population variability of seed production and mean seed mass during four consecutive years. Reproductive traits were highly variable between trees and between years. The high narrow sense heritability and evolvability estimated underline the important genetic effect on the variability in seed production and mean seed mass. Despite a large variability over years, reproductive traits show significant genetic correlation between years. Furthermore, for the first time in forest tree species, quantitative trait loci (QTLs) associated with seed production and mean mass of a seed have been identified. While it is commonly assumed and observed that fitness-traits have low narrow sense heritabilities, our findings show that reproduction-related traits may undergo evolutionary changes under selective pressure and may be determinant for tree adaptation.

Oaks are an important part of our natural and cultural heritage. Not only are they ubiquitous in our most common landscapes 1 but they have also supplied human societies with invaluable services, including food and shelter, since prehistoric times 2 . With 450 species spread throughout Asia, Europe and America 3 , oaks constitute a critical global renewable resource. The longevity of oaks (several hundred years) probably underlies their emblematic cultural and historical importance. Such long-lived sessile organisms must persist in the face of a wide range of abiotic and biotic threats over their lifespans. We investigated the genomic features associated with such a long lifespan by sequencing, assembling and annotating the oak genome. We then used the growing number of whole-genome sequences for plants (including tree and herbaceous species) to investigate the parallel evolution of genomic characteristics potentially underpinning tree longevity. A further consequence of the long lifespan of trees is their accumulation of somatic mutations during mitotic divisions of stem cells present in the shoot apical meristems. Empirical 4 and modelling 5 approaches have shown that intra-organismal genetic heterogeneity can be selected for 6 and provides direct fitness benefits in the arms race with short-lived pests and pathogens through a patchwork of intra-organismal phenotypes 7 . However, there is no clear proof that large-statured trees consist of a genetic mosaic of clonally distinct cell lineages within and between branches. Through this case study of oak, we demonstrate the accumulation and transmission of somatic mutations and the expansion of disease-resistance gene families in trees.

Historical trajectories of tree species during the late Quaternary have been well reconstructed through genetic and palaeobotanical studies. However, many congeneric tree species are interfertile, and the timing and contribution of introgression to species divergence during their evolutionary history remains largely unknown. We quantified past and current gene flow events between four morphologically divergent oak species (Quercus petraea, Q. robur, Q. pyrenaica, Q. pubescens), by two independent inference methods: diffusion approximation to the joint frequency spectrum (∂a∂i) and approximate Bayesian computation (ABC). For each pair of species, alternative scenarios of speciation allowing gene flow over different timescales were evaluated. Analyses of 3524 single nucleotide polymorphisms (SNPs) randomly distributed in the genome, showed that these species evolved in complete isolation for most of their history, but recently came into secondary contact, probably facilitated by the most recent period of postglacial warming. We demonstrated that: there was sufficient genetic differentiation before secondary contact for the accumulation of barriers to gene flow; and current European white oak genomes are a mosaic of genes that have crossed species boundaries and genes impermeable to gene flow., (© 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.)

Testing how populations are locally adapted and predicting their response to their future environment is of key importance in view of climate change. Landscape genomics is a powerful approach to investigate genes and environmental factors involved in local adaptation. In a pooled amplicon sequencing approach of 94 genes in 71 populations, we tested whether >3500 single nucleotide polymorphisms (SNPs) in the three most common oak species in Switzerland (Quercus petraea, Q. pubescens, Q. robur) show an association with abiotic factors related to local topography, historical climate and soil characteristics. In the analysis including all species, the most frequently associated environmental factors were those best describing the habitats of the species. In the species-specific analyses, the most important environmental factors and associated SNPs greatly differed among species. However, we identified one SNP and seven genes that were associated with the same environmental factor across all species. We finally used regressions of allele frequencies of the most strongly associated SNPs along environmental gradients to predict the risk of nonadaptedness (RONA), which represents the average change in allele frequency at climate-associated loci theoretically required to match future climatic conditions. RONA is considerable for some populations and species (up to 48% in single populations) and strongly differs among species. Given the long generation time of oaks, some of the required allele frequency changes might not be realistic to achieve based on standing genetic variation. Hence, future adaptedness requires gene flow or planting of individuals carrying beneficial alleles from habitats currently matching future climatic conditions., (© 2016 John Wiley & Sons Ltd.)

We developed the densest single-nucleotide polymorphism (SNP)-based linkage genetic map to date for the genus Quercus An 8k gene-based SNP array was used to genotype more than 1,000 full-sibs from two intraspecific and two interspecific full-sib families of Quercus petraea and Quercus robur A high degree of collinearity was observed between the eight parental maps of the two species. A composite map was then established with 4,261 SNP markers spanning 742 cM over the 12 linkage groups (LGs) of the oak genome. Nine genomic regions from six LGs displayed highly significant distortions of segregation. Two main hypotheses concerning the mechanisms underlying segregation distortion are discussed: genetic load vs. reproductive barriers. Our findings suggest a predominance of pre-zygotic to post-zygotic barriers., (© The Author 2016. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.)

The 1.5 Gbp/2C genome of pedunculate oak (Quercus robur) has been sequenced. A strategy was established for dealing with the challenges imposed by the sequencing of such a large, complex and highly heterozygous genome by a whole-genome shotgun (WGS) approach, without the use of costly and time-consuming methods, such as fosmid or BAC clone-based hierarchical sequencing methods. The sequencing strategy combined short and long reads. Over 49 million reads provided by Roche 454 GS-FLX technology were assembled into contigs and combined with shorter Illumina sequence reads from paired-end and mate-pair libraries of different insert sizes, to build scaffolds. Errors were corrected and gaps filled with Illumina paired-end reads and contaminants detected, resulting in a total of 17,910 scaffolds (>2 kb) corresponding to 1.34 Gb. Fifty per cent of the assembly was accounted for by 1468 scaffolds (N50 of 260 kb). Initial comparison with the phylogenetically related Prunus persica gene model indicated that genes for 84.6% of the proteins present in peach (mean protein coverage of 90.5%) were present in our assembly. The second and third steps in this project are genome annotation and the assignment of scaffolds to the oak genetic linkage map. In accordance with the Bermuda and Fort Lauderdale agreements and the more recent Toronto Statement, the oak genome data have been released into public sequence repositories in advance of publication. In this presubmission paper, the oak genome consortium describes its principal lines of work and future directions for analyses of the nature, function and evolution of the oak genome., (© 2015 John Wiley & Sons Ltd.)

Background: Genetic markers and linkage mapping are basic prerequisites for comparative genetic analyses, QTL detection and map-based cloning. A large number of mapping populations have been developed for oak, but few gene-based markers are available for constructing integrated genetic linkage maps and comparing gene order and QTL location across related species., Results: We developed a set of 573 expressed sequence tag-derived simple sequence repeats (EST-SSRs) and located 397 markers (EST-SSRs and genomic SSRs) on the 12 oak chromosomes (2n = 2x = 24) on the basis of Mendelian segregation patterns in 5 full-sib mapping pedigrees of two species: Quercus robur (pedunculate oak) and Quercus petraea (sessile oak). Consensus maps for the two species were constructed and aligned. They showed a high degree of macrosynteny between these two sympatric European oaks. We assessed the transferability of EST-SSRs to other Fagaceae genera and a subset of these markers was mapped in Castanea sativa, the European chestnut. Reasonably high levels of macrosynteny were observed between oak and chestnut. We also obtained diversity statistics for a subset of EST-SSRs, to support further population genetic analyses with gene-based markers. Finally, based on the orthologous relationships between the oak, Arabidopsis, grape, poplar, Medicago, and soybean genomes and the paralogous relationships between the 12 oak chromosomes, we propose an evolutionary scenario of the 12 oak chromosomes from the eudicot ancestral karyotype., Conclusions: This study provides map locations for a large set of EST-SSRs in two oak species of recognized biological importance in natural ecosystems. This first step toward the construction of a gene-based linkage map will facilitate the assignment of future genome scaffolds to pseudo-chromosomes. This study also provides an indication of the potential utility of new gene-based markers for population genetics and comparative mapping within and beyond the Fagaceae.

Pedunculate (Quercus robur L.) and sessile oak (Quercus petreae Matt. Liebl.) are closely related species with a widely sympatric distribution in Europe. These two oak species are also known to display different ecological features, particularly related to their adaptation to soil waterlogging. Pedunculate oak grows in humid areas and can withstand high moisture content of the soil, whereas sessile oak requires drier soil with better drainage. The main goal of this study was to explore the role of gene expression contributing to differences in terms of waterlogging tolerance between these two species. We implemented a series of experiments aimed at evaluating whether differentially expressed genes between species are associated with their ecological preferences and underlie adaptive genetic divergence. Rooted cuttings of both species were grown in hydroponic conditions and subjected to gradual root hypoxia. White roots were sampled after 6, 12, 24 and 48 h. Real-time polymerase chain reaction (qPCR) was first used to monitor the expression of 10 known waterlogging-responsive genes, to identify discriminating sampling time points along the kinetics of hypoxia. Secondly, four subtractive suppressive hybridization libraries (sessile vs. pedunculate, pedunculate vs. sessile for early and late responses) were generated to isolate differentially expressed genes between species. A total of 2160 high-quality expressed sequence tags were obtained and annotated, and a subset of 45 genes were selected for qPCR analysis in a second independent factorial experimental design applying two stress durations per two species. Significant differences of gene expression between pedunculate and sessile oaks were detected, suggesting species-specific molecular strategies to respond to hypoxia. This study revealed that the ability of pedunculate oak to maintain glycolysis and fermentation under hypoxic conditions may help explain its tolerance to waterlogging.

Background: One of the key goals of oak genomics research is to identify genes of adaptive significance. This information may help to improve the conservation of adaptive genetic variation and the management of forests to increase their health and productivity. Deep-coverage large-insert genomic libraries are a crucial tool for attaining this objective. We report herein the construction of a BAC library for Quercus robur, its characterization and an analysis of BAC end sequences., Results: The EcoRI library generated consisted of 92,160 clones, 7% of which had no insert. Levels of chloroplast and mitochondrial contamination were below 3% and 1%, respectively. Mean clone insert size was estimated at 135 kb. The library represents 12 haploid genome equivalents and, the likelihood of finding a particular oak sequence of interest is greater than 99%. Genome coverage was confirmed by PCR screening of the library with 60 unique genetic loci sampled from the genetic linkage map. In total, about 20,000 high-quality BAC end sequences (BESs) were generated by sequencing 15,000 clones. Roughly 5.88% of the combined BAC end sequence length corresponded to known retroelements while ab initio repeat detection methods identified 41 additional repeats. Collectively, characterized and novel repeats account for roughly 8.94% of the genome. Further analysis of the BESs revealed 1,823 putative genes suggesting at least 29,340 genes in the oak genome. BESs were aligned with the genome sequences of Arabidopsis thaliana, Vitis vinifera and Populus trichocarpa. One putative collinear microsyntenic region encoding an alcohol acyl transferase protein was observed between oak and chromosome 2 of V. vinifera., Conclusions: This BAC library provides a new resource for genomic studies, including SSR marker development, physical mapping, comparative genomics and genome sequencing. BES analysis provided insight into the structure of the oak genome. These sequences will be used in the assembly of a future genome sequence for oak.

Background: Expressed Sequence Tags (ESTs) are a source of simple sequence repeats (SSRs) that can be used to develop molecular markers for genetic studies. The availability of ESTs for Quercus robur and Quercus petraea provided a unique opportunity to develop microsatellite markers to accelerate research aimed at studying adaptation of these long-lived species to their environment. As a first step toward the construction of a SSR-based linkage map of oak for quantitative trait locus (QTL) mapping, we describe the mining and survey of EST-SSRs as well as a fast and cost-effective approach (bin mapping) to assign these markers to an approximate map position. We also compared the level of polymorphism between genomic and EST-derived SSRs and address the transferability of EST-SSRs in Castanea sativa (chestnut)., Results: A catalogue of 103,000 Sanger ESTs was assembled into 28,024 unigenes from which 18.6% presented one or more SSR motifs. More than 42% of these SSRs corresponded to trinucleotides. Primer pairs were designed for 748 putative unigenes. Overall 37.7% (283) were found to amplify a single polymorphic locus in a reference full-sib pedigree of Quercus robur. The usefulness of these loci for establishing a genetic map was assessed using a bin mapping approach. Bin maps were constructed for the male and female parental tree for which framework linkage maps based on AFLP markers were available. The bin set consisting of 14 highly informative offspring selected based on the number and position of crossover sites. The female and male maps comprised 44 and 37 bins, with an average bin length of 16.5 cM and 20.99 cM, respectively. A total of 256 EST-SSRs were assigned to bins and their map position was further validated by linkage mapping. EST-SSRs were found to be less polymorphic than genomic SSRs, but their transferability rate to chestnut, a phylogenetically related species to oak, was higher., Conclusion: We have generated a bin map for oak comprising 256 EST-SSRs. This resource constitutes a first step toward the establishment of a gene-based map for this genus that will facilitate the dissection of QTLs affecting complex traits of ecological importance.

Nucleotide diversity was assessed within nine candidate genes (in total 4.6 kb) for the time of bud burst in nine sessile oak (Quercus petraea) populations distributed in central and northern Europe. The sampled populations were selected on the basis of their contrasting time of bud burst observed in common garden experiments (provenance tests). The candidate genes were selected according to their expression profiles during the transition from quiescent to developing buds and/or their functional role in model plants. The overall nucleotide diversity was large (π(tot)=6.15 × 10(-3); π(silent)=11.2 × 10(-3)), but population differentiation was not larger than for microsatellites. No outlier single-nucleotide polymorphism (SNP), departing from neutral expectation, was found among the total of 125 SNPs. These results contrasted markedly with the significant associations that were observed between the candidate genes and bud burst in segregating populations. Quantitative trait loci (QTLs) for bud burst were identified for 13 year*site seasonal observations in a cloned mapping pedigree. Nineteen QTLs were detected, and QTLs located on linkage groups 2, 5 and 9 contributed repeatedly to more than 12% of the phenotypic variation of the trait. Eight genes were polymorphic in the two parents of the pedigree and could be mapped on the existing genetic map. Five of them located within the confidence intervals of QTLs for bud burst. Interestingly, four of them located within the three QTLs exhibiting the largest contributions to bud burst.

Interspecific differentiation values (G(ST)) between two closely related oak species (Quercus petraea and Q. robur) were compiled across different studies with the aim to explore the distribution of differentiation at the genome level. The study was based on a total set of 389 markers (isozymes, AFLPs, SCARs, microsatellites, and SNPs) for which allelic frequencies were estimated in pairs of populations sampled throughout the sympatric distribution of the two species. The overall distribution of G(ST) values followed an L-shaped curve with most markers exhibiting low species differentiation (G(ST) < 0.01) and only a few loci reaching >10% levels. Twelve percent of the loci exhibited significant G(ST) deviations to neutral expectations, suggesting that selection contributed to species divergence. Coding regions expressed higher differentiation than noncoding regions. Among the 389 markers, 158 could be mapped on the 12 linkage groups of the existing Q. robur genetic map. Outlier loci with large G(ST) values were distributed over 9 linkage groups. One cluster of three outlier loci was found within 0.51 cM; but significant autocorrelation of G(ST) was observed at distances <2 cM. The size and distribution of genomic regions involved in species divergence are discussed in reference to hitchhiking effects and disruptive selection.

Genetic variation of bud burst and early growth components was estimated in a full-sib family of Quercus robur L. comprising 278 offspring. The full sibs were vegetatively propagated, and phenotypic assessments were made in three field tests. This two-generation pedigree was also used to construct a genetic linkage map (12 linkage groups, 128 markers) and locate quantitative trait loci (QTLs) controlling bud burst and growth components. In each field test, the date of bud burst extended over a period of 20 days from the earliest to the latest clone. Bud burst exhibited higher heritability (0.15-0.51) than growth components (0.04-0.23) and also higher correlations across field tests. Over the three tests there were 32 independent detected QTLs ( P