National Clonal Germplasm Repository

The International Strawberry Sequencing Consortium reports the draft genome of the woodland strawberry (Fragaria vesca). The genome of this diploid species should serve as a reference genome for the Fragaria genus, as the cultivated strawberry (Fragaria × ananassa) is an octoploid where F. vesca is predicted to be a subgenome donor. The woodland strawberry, Fragaria vesca (2n = 2x = 14), is a versatile experimental plant system. This diminutive herbaceous perennial has a small genome (240 Mb), is amenable to genetic transformation and shares substantial sequence identity with the cultivated strawberry (Fragaria × ananassa) and other economically important rosaceous plants. Here we report the draft F. vesca genome, which was sequenced to ×39 coverage using second-generation technology, assembled de novo and then anchored to the genetic linkage map into seven pseudochromosomes. This diploid strawberry sequence lacks the large genome duplications seen in other rosids. Gene prediction modeling identified 34,809 genes, with most being supported by transcriptome mapping. Genes critical to valuable horticultural traits including flavor, nutritional value and flowering time were identified. Macrosyntenic relationships between Fragaria and Prunus predict a hypothetical ancestral Rosaceae genome that had nine chromosomes. New phylogenetic analysis of 154 protein-coding genes suggests that assignment of Populus to Malvidae, rather than Fabidae, is warranted.

The grape is one of the earliest domesticated fruit crops and, since antiquity, it has been widely cultivated and prized for its fruit and wine. Here, we characterize genome-wide patterns of genetic variation in over 1,000 samples of the domesticated grape, Vitis vinifera subsp. vinifera, and its wild relative, V. vinifera subsp. sylvestris from the US Department of Agriculture grape germplasm collection. We find support for a Near East origin of vinifera and present evidence of introgression from local sylvestris as the grape moved into Europe. High levels of genetic diversity and rapid linkage disequilibrium (LD) decay have been maintained in vinifera, which is consistent with a weak domestication bottleneck followed by thousands of years of widespread vegetative propagation. The considerable genetic diversity within vinifera, however, is contained within a complex network of close pedigree relationships that has been generated by crosses among elite cultivars. We show that first-degree relationships are rare between wine and table grapes and among grapes from geographically distant regions. Our results suggest that although substantial genetic diversity has been maintained in the grape subsequent to domestication, there has been a limited exploration of this diversity. We propose that the adoption of vegetative propagation was a double-edged sword: Although it provided a benefit by ensuring true breeding cultivars, it also discouraged the generation of unique cultivars through crosses. The grape currently faces severe pathogen pressures, and the long-term sustainability of the grape and wine industries will rely on the exploitation of the grape's tremendous natural genetic diversity.

Abstract The domestication of the Eurasian grape ( Vitis vinifera ssp. sativa ) from its wild ancestor ( Vitis vinifera ssp. sylvestris ) has long been claimed to have occurred in Transcaucasia where its greatest genetic diversity is found and where very early archaeological evidence, including grape pips and artefacts of a ‘wine culture’, have been excavated. Whether from Transcaucasia or the nearby Taurus or Zagros Mountains, it is hypothesized that this wine culture spread southwards and eventually westwards around the Mediterranean basin, together with the transplantation of cultivated grape cuttings. However, the existence of morphological differentiation between cultivars from eastern and western ends of the modern distribution of the Eurasian grape suggests the existence of different genetic contribution from local sylvestris populations or multilocal selection and domestication of sylvestris genotypes. To tackle this issue, we analysed chlorotype variation and distribution in 1201 samples of sylvestris and sativa genotypes from the whole area of the species’ distribution and studied their genetic relationships. The results suggest the existence of at least two important origins for the cultivated germplasm, one in the Near East and another in the western Mediterranean region, the latter of which gave rise to many of the current Western European cultivars. Indeed, over 70% of the Iberian Peninsula cultivars display chlorotypes that are only compatible with their having derived from western sylvestris populations.

BACKGROUND: Pear (Pyrus) is a globally grown fruit, with thousands of cultivars in five domesticated species and dozens of wild species. However, little is known about the evolutionary history of these pear species and what has contributed to the distinct phenotypic traits between Asian pears and European pears. RESULTS: We report the genome resequencing of 113 pear accessions from worldwide collections, representing both cultivated and wild pear species. Based on 18,302,883 identified SNPs, we conduct phylogenetics, population structure, gene flow, and selective sweep analyses. Furthermore, we propose a model for the divergence, dissemination, and independent domestication of Asian and European pears in which pear, after originating in southwest China and then being disseminated throughout central Asia, has eventually spread to western Asia, and then on to Europe. We find evidence for rapid evolution and balancing selection for S-RNase genes that have contributed to the maintenance of self-incompatibility, thus promoting outcrossing and accounting for pear genome diversity across the Eurasian continent. In addition, separate selective sweep signatures between Asian pears and European pears, combined with co-localized QTLs and differentially expressed genes, underline distinct phenotypic fruit traits, including flesh texture, sugar, acidity, aroma, and stone cells. CONCLUSIONS: This study provides further clarification of the evolutionary history of pear along with independent domestication of Asian and European pears. Furthermore, it provides substantive and valuable genomic resources that will significantly advance pear improvement and molecular breeding efforts.

As high-throughput genetic marker screening systems are essential for a range of genetics studies and plant breeding applications, the International RosBREED SNP Consortium (IRSC) has utilized the Illumina Infinium® II system to develop a medium- to high-throughput SNP screening tool for genome-wide evaluation of allelic variation in apple (Malus×domestica) breeding germplasm. For genome-wide SNP discovery, 27 apple cultivars were chosen to represent worldwide breeding germplasm and re-sequenced at low coverage with the Illumina Genome Analyzer II. Following alignment of these sequences to the whole genome sequence of 'Golden Delicious', SNPs were identified using SoapSNP. A total of 2,113,120 SNPs were detected, corresponding to one SNP to every 288 bp of the genome. The Illumina GoldenGate® assay was then used to validate a subset of 144 SNPs with a range of characteristics, using a set of 160 apple accessions. This validation assay enabled fine-tuning of the final subset of SNPs for the Illumina Infinium® II system. The set of stringent filtering criteria developed allowed choice of a set of SNPs that not only exhibited an even distribution across the apple genome and a range of minor allele frequencies to ensure utility across germplasm, but also were located in putative exonic regions to maximize genotyping success rate. A total of 7867 apple SNPs was established for the IRSC apple 8K SNP array v1, of which 5554 were polymorphic after evaluation in segregating families and a germplasm collection. This publicly available genomics resource will provide an unprecedented resolution of SNP haplotypes, which will enable marker-locus-trait association discovery, description of the genetic architecture of quantitative traits, investigation of genetic variation (neutral and functional), and genomic selection in apple.

222 cultivated (Vitis vinifera) and 22 wild (V. vinifera ssp. sylvestris) grape accessions were analysed for genetic diversity and differentiation at eight microsatellite loci. A total of 94 alleles were detected, with extensive polymorphism among the accessions. Multivariate relationships among accessions revealed 16 genetic groups structured into three clusters, supporting the classical eco-geographic grouping of grape cultivars: occidentalis, pontica and orientalis. French cultivars appeared to be distinct and showed close affinity to the wild progenitor, ssp. sylvestris from south-western France (Pyrenees) and Tunisia, probably reflecting the origin and domestication history of many of the old wine cultivars from France. There was appreciable level of differentiation between table and wine grape cultivars, and the Muscat types were somewhat distinct within the wine grapes. Contingency chi2 analysis indicated significant heterogeneity in allele frequencies among groups at all loci. The observed heterozygosities for different groups ranged from 0.625 to 0.9 with an overall average of 0.771. Genetic relationships among groups suggested hierarchical differentiation within cultivated grape. The gene diversity analysis indicated narrow divergence among groups and that most variation was found within groups (approximately 85%). Partitioning of diversity suggested that the remaining variation is somewhat structured hierarchically at different levels of differentiation. The overall organization of genetic diversity suggests that the germplasm of cultivated grape represents a single complex gene pool and that its structure is determined by strong artificial selection and a vegetative mode of reproduction.

In this paper, a broad overview is provide<;l for the fruit known as the pomegranate (Pu~ica granatum). The pomegranate has a deep association with the cultures of the Mediterranean region and Near East, where it is savored as a delicacy and is an important dietary component, revered in symbolism, and greatly appreciated for its medicinal properties. It is strange that a horticultural icon of such importance has been largely relegated to an ornamental role in the United States and much of Europe. Recent trends indicate that the health-giving and flavorfilled properties of these fruits may soon reverse this oversight.

The Persian walnut (Juglans regia L.), a diploid species native to the mountainous regions of Central Asia, is the major walnut species cultivated for nut production and is one of the most widespread tree nut species in the world. The high nutritional value of J. regia nuts is associated with a rich array of polyphenolic compounds, whose complete biosynthetic pathways are still unknown. A J. regia genome sequence was obtained from the cultivar 'Chandler' to discover target genes and additional unknown genes. The 667-Mbp genome was assembled using two different methods (SOAPdenovo2 and MaSuRCA), with an N50 scaffold size of 464 955 bp (based on a genome size of 606 Mbp), 221 640 contigs and a GC content of 37%. Annotation with MAKER-P and other genomic resources yielded 32 498 gene models. Previous studies in walnut relying on tissue-specific methods have only identified a single polyphenol oxidase (PPO) gene (JrPPO1). Enabled by the J. regia genome sequence, a second homolog of PPO (JrPPO2) was discovered. In addition, about 130 genes in the large gallate 1-β-glucosyltransferase (GGT) superfamily were detected. Specifically, two genes, JrGGT1 and JrGGT2, were significantly homologous to the GGT from Quercus robur (QrGGT), which is involved in the synthesis of 1-O-galloyl-β-d-glucose, a precursor for the synthesis of hydrolysable tannins. The reference genome for J. regia provides meaningful insight into the complex pathways required for the synthesis of polyphenols. The walnut genome sequence provides important tools and methods to accelerate breeding and to facilitate the genetic dissection of complex traits.

This chapter contains sections titled: Introduction Variability and Genetic Resources Plant Morphology and Development Horticulture Postharvest Physiology and Handling Genetics and Breeding Human Nutrition and Health Conclusion Acknowledgments Literature Cited

Terroir, the unique interaction between genotype, environment, and culture, is highly refined in domesticated grape (Vitis vinifera). Toward cultivating terroir, the science of ampelography tried to distinguish thousands of grape cultivars without the aid of genetics. This led to sophisticated phenotypic analyses of natural variation in grape leaves, which within a palmate-lobed framework exhibit diverse patterns of blade outgrowth, hirsuteness, and venation patterning. Here, we provide a morphometric analysis of more than 1,200 grape accessions. Elliptical Fourier descriptors provide a global analysis of leaf outlines and lobe positioning, while a Procrustes analysis quantitatively describes venation patterning. Correlation with previous ampelography suggests an important genetic component, which we confirm with estimates of heritability. We further use RNA-Seq of mutant varieties and perform a genome-wide association study to explore the genetic basis of leaf shape. Meta-analysis reveals a relationship between leaf morphology and hirsuteness, traits known to correlate with climate in the fossil record and extant species. Together, our data demonstrate a genetic basis for the intricate diversity present in grape leaves. We discuss the possibility of using grape leaves as a breeding target to preserve terroir in the face of anticipated climate change, a major problem facing viticulture.

Although a large number of single nucleotide polymorphism (SNP) markers covering the entire genome are needed to enable molecular breeding efforts such as genome wide association studies, fine mapping, genomic selection and marker-assisted selection in peach [Prunus persica (L.) Batsch] and related Prunus species, only a limited number of genetic markers, including simple sequence repeats (SSRs), have been available to date. To address this need, an international consortium (The International Peach SNP Consortium; IPSC) has pursued a coordinated effort to perform genome-scale SNP discovery in peach using next generation sequencing platforms to develop and characterize a high-throughput Illumina Infinium® SNP genotyping array platform. We performed whole genome re-sequencing of 56 peach breeding accessions using the Illumina and Roche/454 sequencing technologies. Polymorphism detection algorithms identified a total of 1,022,354 SNPs. Validation with the Illumina GoldenGate® assay was performed on a subset of the predicted SNPs, verifying ∼75% of genic (exonic and intronic) SNPs, whereas only about a third of intergenic SNPs were verified. Conservative filtering was applied to arrive at a set of 8,144 SNPs that were included on the IPSC peach SNP array v1, distributed over all eight peach chromosomes with an average spacing of 26.7 kb between SNPs. Use of this platform to screen a total of 709 accessions of peach in two separate evaluation panels identified a total of 6,869 (84.3%) polymorphic SNPs.The almost 7,000 SNPs verified as polymorphic through extensive empirical evaluation represent an excellent source of markers for future studies in genetic relatedness, genetic mapping, and dissecting the genetic architecture of complex agricultural traits. The IPSC peach SNP array v1 is commercially available and we expect that it will be used worldwide for genetic studies in peach and related stone fruit and nut species.

BACKGROUND: Rapid development of highly saturated genetic maps aids molecular breeding, which can accelerate gain per breeding cycle in woody perennial plants such as Rubus idaeus (red raspberry). Recently, robust genotyping methods based on high-throughput sequencing were developed, which provide high marker density, but result in some genotype errors and a large number of missing genotype values. Imputation can reduce the number of missing values and can correct genotyping errors, but current methods of imputation require a reference genome and thus are not an option for most species. RESULTS: Genotyping by Sequencing (GBS) was used to produce highly saturated maps for a R. idaeus pseudo-testcross progeny. While low coverage and high variance in sequencing resulted in a large number of missing values for some individuals, a novel method of imputation based on maximum likelihood marker ordering from initial marker segregation overcame the challenge of missing values, and made map construction computationally tractable. The two resulting parental maps contained 4521 and 2391 molecular markers spanning 462.7 and 376.6 cM respectively over seven linkage groups. Detection of precise genomic regions with segregation distortion was possible because of map saturation. Microsatellites (SSRs) linked these results to published maps for cross-validation and map comparison. CONCLUSIONS: GBS together with genome-independent imputation provides a rapid method for genetic map construction in any pseudo-testcross progeny. Our method of imputation estimates the correct genotype call of missing values and corrects genotyping errors that lead to inflated map size and reduced precision in marker placement. Comparison of SSRs to published R. idaeus maps showed that the linkage maps constructed with GBS and our method of imputation were robust, and marker positioning reliable. The high marker density allowed identification of genomic regions with segregation distortion in R. idaeus, which may help to identify deleterious alleles that are the basis of inbreeding depression in the species.

Abstract Heat treatments have been used to control fungal diseases and insect infestation of fruit for many years. However, with the development of effective fungicides and insecticides, especially fumigants, which could be applied cheaply and easily, interest in heat treatments waned. Stringent short- and long-term safety studies imposed by the U.S. Environmental Protection Agency has made retention of registration for many agricultural chemicals increasingly difficult. Some of the fumigants, such as ethylene dibromide, which were developed and used to control insects during the past 20 to 30 years, are no longer registered; others may lose registration in the future. These regulatory restrictions also increase the cost of developing new chemical fumigants and, therefore, interest in heat disinfestation has been revived (5).

Abstract Phylogenetic relationships among 48 species of Prunus were investigated by means of maximum parsimony analysis of sequence polymorphism from nuclear ITS and chloroplast trnL-trnF spacer DNA. Several genera that have been implicated as close relatives of Prunus in previous studies were utilized as outgroups. The objective of this study was to reconstruct the phylogeny of Prunus with the purpose of reviewing previously described taxonomic relationships and providing a basis for studies of morphological evolution in the genus. Variability and homoplasy are higher in ITS than in trnL-trnF. Parsimony analysis of both markers, separately and in combination, supported a close relationship among Exochorda, Oemleria, and Prinsepia but did not support a sister relationship of this clade with Prunus. Prunus is monophyletic and divided into two groups. The first is a strongly supported clade composed of subgenera Padus, Laurocerasus, and Cerasus. None of these subgenera is monophyletic and the trait used to characterize Padus and Laurocerasus, flowers arranged in racemes, can be more broadly interpreted as a synapomorphy for the clade. The second group included the subgenera Amygdalus and Prunus, and sections Microcerasus (subgenus Cerasus), and Penarmeniaca, which has never been assigned to a subgenus. The exclusively North American subgenus Emplectocladus may be an early diverging taxon sister to the second group. Sections Prunus and Armeniaca are monophyletic with low bootstrap support. Section Penarmeniaca, from southwestern US, and P. besseyi (section Microcerasus) are sister taxa. A Eurasian origin of Prunus is supported. Communicating Editor: Richard Jensen

BACKGROUND: The mountainous region between the Caucasus and China is considered to be the center of domestication for grapevine. Despite the importance of Central Asia in the history of grape growing, information about the extent and distribution of grape genetic variation in this region is limited in comparison to wild and cultivated grapevines from around the Mediterranean basin. The principal goal of this work was to survey the genetic diversity and relationships among wild and cultivated grape germplasm from the Caucasus, Central Asia, and the Mediterranean basin collectively to understand gene flow, possible domestication events and adaptive introgression. RESULTS: analysis and assignment tests. All of these analyses identified three genetic groups: G1, wild accessions from Croatia, France, Italy and Spain; G2, wild accessions from Armenia, Azerbaijan and Georgia; and G3, cultivars from Spain, France, Italy, Georgia, Iran, Pakistan and Turkmenistan, which included a small group of wild accessions from Georgia and Croatia. Wild accessions from Georgia clustered with cultivated grape from the same area (proles pontica), but also with Western Europe (proles occidentalis), supporting Georgia as the ancient center of grapevine domestication. In addition, cluster analysis indicated that Western European wild grapes grouped with cultivated grapes from the same area, suggesting that the cultivated proles occidentalis contributed more to the early development of wine grapes than the wild vines from Eastern Europe. CONCLUSIONS: The analysis of genetic relationships among the tested genotypes provided evidence of genetic relationships between wild and cultivated accessions in the Mediterranean basin and Central Asia. The genetic structure indicated a considerable amount of gene flow, which limited the differentiation between the two subspecies. The results also indicated that grapes with mixed ancestry occur in the regions where wild grapevines were domesticated.

Summary For over a century, white pine blister rust ( Cronartium ribicola ) has linked white pines ( Strobus ) with currants and gooseberries ( Ribes ) in a complex and serious disease epidemic in Asia, Europe, and North America. Because of ongoing changes in climate, societal demands for forests and their amenities, and scientific advances in genetics and proteomics, our current understanding and management of the white pine blister rust pathosystem has become outdated. Here, we present a review and synthesis of international scope on the biology and management of blister rust, white pines, Ribes , and other hosts. In this article, we provide a geographical and historical background, describe the taxonomy and life cycle of the rust, discuss pathology and ecology, and introduce a series of invited papers. These review articles summarize the literature on white pines, Ribes , and blister rust with respect to their status, threats, and management through genetics and silviculture. Although the principal focus is on North America, the different epidemics in Europe and Asia are also described. In the final article, we discuss several of the key observations and conclusions from the preceding review articles and identify prudent actions for research and management of white pine blister rust.

The genus Ficus includes species ranging in number from 600 to more than 1900, with most found in the tropics or subtropics and only a handful with fruits considered edible (reviewed in The cultivated fig, Ficus carica L., (Moraceae), is clearly of greatest importance as a source of human food. The fig fruit has long been associated with horticulture in the Mediterranean region (Zohary and Spiegel-Roy, 1975) and is considered to have been ''first brought into cultivation in southern Arabia'' (Storey, 1975). Wild or ''nearly wild'' figs are reported throughout much of the Middle East and Mediterranean region (De Candolle, 1886). Cultivated figs are reported to have become established across the Mediterranean region 6000 years ago, reaching England by 500 CE Interestingly, the fossil record shows a prehistoric distribution of Ficus carica across southern Europe (De Candolle, 1886).

Black raspberry (Rubus occidentalis) is an important specialty fruit crop in the US Pacific Northwest that can hybridize with the globally commercialized red raspberry (R. idaeus). Here we report a 243 Mb draft genome of black raspberry that will serve as a useful reference for the Rosaceae and Rubus fruit crops (raspberry, blackberry, and their hybrids). The black raspberry genome is largely collinear to the diploid woodland strawberry (Fragaria vesca) with a conserved karyotype and few notable structural rearrangements. Centromeric satellite repeats are widely dispersed across the black raspberry genome, in contrast to the tight association with the centromere observed in most plants. Among the 28 005 predicted protein-coding genes, we identified 290 very recent small-scale gene duplicates enriched for sugar metabolism, fruit development, and anthocyanin related genes which may be related to key agronomic traits during black raspberry domestication. This contrasts patterns of recent duplications in the wild woodland strawberry F. vesca, which show no patterns of enrichment, suggesting gene duplications contributed to domestication traits. Expression profiles from a fruit ripening series and roots exposed to Verticillium dahliae shed insight into fruit development and disease response, respectively. The resources presented here will expedite the development of improved black and red raspberry, blackberry and other Rubus cultivars.

Background: The fragmented nature of most draft plant genomes has hindered downstream gene discovery, trait mapping for breeding, and other functional genomics applications. There is a pressing need to improve or finish draft plant genome assemblies. Findings: Here, we present a chromosome-scale assembly of the black raspberry genome using single-molecule real-time Pacific Biosciences sequencing and high-throughput chromatin conformation capture (Hi-C) genome scaffolding. The updated V3 assembly has a contig N50 of 5.1 Mb, representing an ∼200-fold improvement over the previous Illumina-based version. Each of the 235 contigs was anchored and oriented into seven chromosomes, correcting several major misassemblies. Black raspberry V3 contains 47 Mb of new sequences including large pericentromeric regions and thousands of previously unannotated protein-coding genes. Among the new genes are hundreds of expanded tandem gene arrays that were collapsed in the Illumina-based assembly. Detailed comparative genomics with the high-quality V4 woodland strawberry genome (Fragaria vesca) revealed near-perfect 1:1 synteny with dramatic divergence in tandem gene array composition. Lineage-specific tandem gene arrays in black raspberry are related to agronomic traits such as disease resistance and secondary metabolite biosynthesis. Conclusions: The improved resolution of tandem gene arrays highlights the need to reassemble these highly complex and biologically important regions in draft plant genomes. The updated, high-quality black raspberry reference genome will be useful for comparative genomics across the horticulturally important Rosaceae family and enable the development of marker assisted breeding in Rubus.

BACKGROUND: During evolution, plants and other organisms have developed a diversity of chemical defences, leading to the evolution of various groups of specialized metabolites selected for their endogenous biological function. A correlation between phylogeny and biosynthetic pathways could offer a predictive approach enabling more efficient selection of plants for the development of traditional medicine and lead discovery. However, this relationship has rarely been rigorously tested and the potential predictive power is consequently unknown. RESULTS: We produced a phylogenetic hypothesis for the medicinally important plant subfamily Amaryllidoideae (Amaryllidaceae) based on parsimony and Bayesian analysis of nuclear, plastid, and mitochondrial DNA sequences of over 100 species. We tested if alkaloid diversity and activity in bioassays related to the central nervous system are significantly correlated with phylogeny and found evidence for a significant phylogenetic signal in these traits, although the effect is not strong. CONCLUSIONS: Several genera are non-monophyletic emphasizing the importance of using phylogeny for interpretation of character distribution. Alkaloid diversity and in vitro inhibition of acetylcholinesterase (AChE) and binding to the serotonin reuptake transporter (SERT) are significantly correlated with phylogeny. This has implications for the use of phylogenies to interpret chemical evolution and biosynthetic pathways, to select candidate taxa for lead discovery, and to make recommendations for policies regarding traditional use and conservation priorities.