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Heterosis was the catalyst for the domestication of cultivated strawberry (Fragaria × ananassa), an interspecific hybrid species that originated in the 1700s. The hybrid origin was discovered because the phenotypes of spontaneous hybrids transgressed those of their parent species. The transgressions included fruit yield increases and other genetic gains in the twentieth century that sparked the global expansion of strawberry production. The importance of heterosis to the agricultural success of the hybrid species, however, has remained a mystery. Here we show that heterosis has disappeared (become fixed) among improved hybrids within a population (the California population) that has been under long-term selection for increased fruit yield, weight, and firmness. We found that the highest yielding hybrids are among the most highly inbred (59-79%), which seems counterintuitive for a highly heterozygous, outbreeder carrying heavy genetic loads. Although faint remnants of heterosis were discovered, the between-parent allele frequency differences and dispersed favorable dominant alleles necessary for heterosis have decreased nearly genome-wide within the California population. Conversely, heterosis was prevalent and significant among wide hybrids, especially for fruit count, a significant driver of genetic gains for fruit yield. We attributed the disappearance (fixation) of heterosis within the California population to increased homozygosity of favorable dominant alleles and inbreeding associated with selection, random genetic drift, and selective sweeps. Despite historical inbreeding, the highest yielding hybrids reported to-date are estimated to be heterozygous for 20,370-44,280 of 97,000-108,000 genes in the octoploid genome, the equivalent of an entire diploid genome or more.

The development of strawberry (Fragaria × ananassa Duchesne ex Rozier) cultivars resistant to Phytophthora crown rot (PhCR), a devastating disease caused by the soil-borne pathogen Phytophthora cactorum (Lebert & Cohn) J. Schröt., has been challenging partly because the resistance phenotypes are quantitative and only moderately heritable. To develop deeper insights into the genetics of resistance and build the foundation for applying genomic selection, a genetically diverse training population was screened for resistance to California isolates of the pathogen. Here we show that genetic gains in breeding for resistance to PhCR have been negligible (3% of the cultivars tested were highly resistant and none surpassed early 20th century cultivars). Narrow-sense genomic heritability for PhCR resistance ranged from 0.41 to 0.75 among training population individuals. Using multivariate genome-wide association studies (GWAS), we identified a large-effect locus (predicted to be RPc2) that explained 43.6-51.6% of the genetic variance, was necessary but not sufficient for resistance, and was associated with calcium channel and other candidate genes with known plant defense functions. The addition of underutilized gene bank resources to our training population doubled additive genetic variance, increased the accuracy of genomic selection, and enabled the discovery of individuals carrying favorable alleles that are either rare or not present in modern cultivars. The incorporation of an RPc2-associated single-nucleotide polymorphism (SNP) as a fixed effect increased genomic prediction accuracy from 0.40 to 0.55. Finally, we show that parent selection using genomic-estimated breeding values, genetic variances, and cross usefulness holds promise for enhancing resistance to PhCR in strawberry.

Flavor is essential to consumer preference of foods and is an increasing focus of plant breeding programs. In fruit crops, identifying genes underlying volatile organic compounds has great promise to accelerate flavor improvement, but polyploidy and heterozygosity in many species have slowed progress. Here we use octoploid cultivated strawberry to demonstrate how genomic heterozygosity, transcriptomic intricacy and fruit metabolomic diversity can be treated as strengths and leveraged to uncover fruit flavor genes and their regulatory elements. Multi-omics datasets were generated including an expression quantitative trait loci map with 196 diverse breeding lines, haplotype-phased genomes of a highly-flavored breeding selection, a genome-wide structural variant map using five haplotypes, and volatile genome-wide association study (GWAS) with > 300 individuals. Overlaying regulatory elements, structural variants and GWAS-linked allele-specific expression of numerous genes to variation in volatile compounds important to flavor. In one example, the functional role of anthranilate synthase alpha subunit 1 in methyl anthranilate biosynthesis was supported via fruit transient gene expression assays. These results demonstrate a framework for flavor gene discovery in fruit crops and a pathway to molecular breeding of cultivars with complex and desirable flavor.

Key messageSeveral Fusarium wilt resistance genes were discovered, genetically and physically mapped, and rapidly deployed via marker-assisted selection to develop cultivars resistant to Fusarium oxysporum f. sp. fragariae, a devastating soil-borne pathogen of strawberry. Fusarium wilt, a soilborne disease caused by Fusarium oxysporum f. sp. fragariae, poses a significant threat to strawberry (Fragaria [Formula: see text] ananassa) production in many parts of the world. This pathogen causes wilting, collapse, and death in susceptible genotypes. We previously identified a dominant gene (FW1) on chromosome 2B that confers resistance to race 1 of the pathogen, and hypothesized that gene-for-gene resistance to Fusarium wilt was widespread in strawberry. To explore this, a genetically diverse collection of heirloom and modern cultivars and octoploid ecotypes were screened for resistance to Fusarium wilt races 1 and 2. Here, we show that resistance to both races is widespread in natural and domesticated populations and that resistance to race 1 is conferred by partially to completely dominant alleles among loci (FW1, FW2, FW3, FW4, and FW5) found on three non-homoeologous chromosomes (1A, 2B, and 6B). The underlying genes have not yet been cloned and functionally characterized; however, plausible candidates were identified that encode pattern recognition receptors or other proteins known to confer gene-for-gene resistance in plants. High-throughput genotyping assays for SNPs in linkage disequilibrium with FW1-FW5 were developed to facilitate marker-assisted selection and accelerate the development of race 1 resistant cultivars. This study laid the foundation for identifying the genes encoded by FW1-FW5, in addition to exploring the genetics of resistance to race 2 and other races of the pathogen, as a precaution to averting a Fusarium wilt pandemic.

Gray mold, a disease of strawberry (Fragaria × ananassa) caused by the ubiquitous necrotroph Botrytis cinerea, renders fruit unmarketable and causes economic losses in the postharvest supply chain. To explore the feasibility of selecting for increased resistance to gray mold, we undertook genetic and genomic prediction studies in strawberry populations segregating for fruit quality and shelf life traits hypothesized to pleiotropically affect susceptibility. As predicted, resistance to gray mold was heritable but quantitative and genetically complex. While every individual was susceptible, the speed of symptom progression and severity differed. Narrow-sense heritability ranged from 0.38 to 0.71 for lesion diameter (LD) and 0.39 to 0.44 for speed of emergence of external mycelium (EM). Even though significant additive genetic variation was observed for LD and EM, the phenotypic ranges were comparatively narrow and genome-wide analyses did not identify any large-effect loci. Genomic selection (GS) accuracy ranged from 0.28 to 0.59 for LD and 0.37 to 0.47 for EM. Additive genetic correlations between fruit quality and gray mold resistance traits were consistent with prevailing hypotheses: LD decreased as titratable acidity increased, whereas EM increased as soluble solid content decreased and firmness increased. We concluded that phenotypic and GS could be effective for reducing LD and increasing EM, especially in long shelf life populations, but that a significant fraction of the genetic variation for resistance to gray mold was caused by the pleiotropic effects of fruit quality traits that differ among market and shelf life classes.

Cultivated strawberry (Fragaria × ananassa) is one of our youngest domesticates, originating in early eighteenth-century Europe from spontaneous hybrids between wild allo-octoploid species (Fragaria chiloensis and Fragaria virginiana). The improvement of horticultural traits by 300 years of breeding has enabled the global expansion of strawberry production. Here, we describe the genomic history of strawberry domestication from the earliest hybrids to modern cultivars. We observed a significant increase in heterozygosity among interspecific hybrids and a decrease in heterozygosity among domesticated descendants of those hybrids. Selective sweeps were found across the genome in early and modern phases of domestication-59-76% of the selectively swept genes originated in the three less dominant ancestral subgenomes. Contrary to the tenet that genetic diversity is limited in cultivated strawberry, we found that the octoploid species harbor massive allelic diversity and that F. × ananassa harbors as much allelic diversity as either wild founder. We identified 41.8 M subgenome-specific DNA variants among resequenced wild and domesticated individuals. Strikingly, 98% of common alleles and 73% of total alleles were shared between wild and domesticated populations. Moreover, genome-wide estimates of nucleotide diversity were virtually identical in F. chiloensis,F. virginiana, and F. × ananassa (π = 0.0059-0.0060). We found, however, that nucleotide diversity and heterozygosity were significantly lower in modern F. × ananassa populations that have experienced significant genetic gains and have produced numerous agriculturally important cultivars.

Shape is a critical element of the visual appeal of strawberry fruit and is influenced by both genetic and non-genetic determinants. Current fruit phenotyping approaches for external characteristics in strawberry often rely on the human eye to make categorical assessments. However, fruit shape is an inherently multi-dimensional, continuously variable trait and not adequately described by a single categorical or quantitative feature. Morphometric approaches enable the study of complex, multi-dimensional forms but are often abstract and difficult to interpret. In this study, we developed a mathematical approach for transforming fruit shape classifications from digital images onto an ordinal scale called the Principal Progression of k Clusters (PPKC). We use these human-recognizable shape categories to select quantitative features extracted from multiple morphometric analyses that are best fit for genetic dissection and analysis. We transformed images of strawberry fruit into human-recognizable categories using unsupervised machine learning, discovered 4 principal shape categories, and inferred progression using PPKC. We extracted 68 quantitative features from digital images of strawberries using a suite of morphometric analyses and multivariate statistical approaches. These analyses defined informative feature sets that effectively captured quantitative differences between shape classes. Classification accuracy ranged from 68% to 99% for the newly created phenotypic variables for describing a shape. Our results demonstrated that strawberry fruit shapes could be robustly quantified, accurately classified, and empirically ordered using image analyses, machine learning, and PPKC. We generated a dictionary of quantitative traits for studying and predicting shape classes and identifying genetic factors underlying phenotypic variability for fruit shape in strawberry. The methods and approaches that we applied in strawberry should apply to other fruits, vegetables, and specialty crops.

BackgroundPopulus trichocarpa is an important forest tree species for the generation of lignocellulosic ethanol. Understanding the genomic basis of biomass production and chemical composition of wood is fundamental in supporting genetic improvement programs. Considerable variation has been observed in this species for complex traits related to growth, phenology, ecophysiology and wood chemistry. Those traits are influenced by both polygenic control and environmental effects, and their genome architecture and regulation are only partially understood. Genome wide association studies (GWAS) represent an approach to advance that aim using thousands of single nucleotide polymorphisms (SNPs). Genotyping using exome capture methodologies represent an efficient approach to identify specific functional regions of genomes underlying phenotypic variation.ResultsWe identified 813 K SNPs, which were utilized for genotyping 461 P. trichocarpa clones, representing 101 provenances collected from Oregon and Washington, and established in California. A GWAS performed on 20 traits, considering single SNP-marker tests identified a variable number of significant SNPs (p-value

Allo-octoploid cultivated strawberry (Fragaria × ananassa) originated through a combination of polyploid and homoploid hybridization, domestication of an interspecific hybrid lineage, and continued admixture of wild species over the last 300 years. While genes appear to flow freely between the octoploid progenitors, the genome structures and diversity of the octoploid species remain poorly understood. The complexity and absence of an octoploid genome frustrated early efforts to study chromosome evolution, resolve subgenomic structure, and develop a single coherent linkage group nomenclature. Here, we show that octoploid Fragaria species harbor millions of subgenome-specific DNA variants. Their diversity was sufficient to distinguish duplicated (homoeologous and paralogous) DNA sequences and develop 50K and 850K SNP genotyping arrays populated with co-dominant, disomic SNP markers distributed throughout the octoploid genome. Whole-genome shotgun genotyping of an interspecific segregating population yielded 1.9M genetically mapped subgenome variants in 5,521 haploblocks spanning 3,394 cM in F. chiloensis subsp. lucida, and 1.6M genetically mapped subgenome variants in 3,179 haploblocks spanning 2,017 cM in F. × ananassa. These studies provide a dense genomic framework of subgenome-specific DNA markers for seamlessly cross-referencing genetic and physical mapping information and unifying existing chromosome nomenclatures. Using comparative genomics, we show that geographically diverse wild octoploids are effectively diploidized, nearly completely collinear, and retain strong macro-synteny with diploid progenitor species. The preservation of genome structure among allo-octoploid taxa is a critical factor in the unique history of garden strawberry, where unimpeded gene flow supported its origin and domestication through repeated cycles of interspecific hybridization.

Conifers are the dominant plant species throughout the high latitude boreal forests as well as some lower latitude temperate forests of North America, Europe, and Asia. As such, they play an integral economic and ecological role across much of the world. This study focused on the characterization of needle transcriptomes from four ecologically important and understudied North American white pines within the Pinus subgenus Strobus The populations of many Strobus species are challenged by native and introduced pathogens, native insects, and abiotic factors. RNA from the needles of western white pine (Pinus monticola), limber pine (Pinus flexilis), whitebark pine (Pinus albicaulis), and sugar pine (Pinus lambertiana) was sampled, Illumina short read sequenced, and de novo assembled. The assembled transcripts and their subsequent structural and functional annotations were processed through custom pipelines to contend with the challenges of non-model organism transcriptome validation. Orthologous gene family analysis of over 58,000 translated transcripts, implemented through Tribe-MCL, estimated the shared and unique gene space among the four species. This revealed 2025 conserved gene families, of which 408 were aligned to estimate levels of divergence and reveal patterns of selection. Specific candidate genes previously associated with drought tolerance and white pine blister rust resistance in conifers were investigated.

Sugar pine (Pinus lambertiana Douglas) is within the subgenus Strobus with an estimated genome size of 31 Gbp. Transcriptomic resources are of particular interest in conifers due to the challenges presented in their megagenomes for gene identification. In this study, we present the first comprehensive survey of the P. lambertiana transcriptome through deep sequencing of a variety of tissue types to generate more than 2.5 billion short reads. Third generation, long reads generated through PacBio Iso-Seq have been included for the first time in conifers to combat the challenges associated with de novo transcriptome assembly. A technology comparison is provided here to contribute to the otherwise scarce comparisons of second and third generation transcriptome sequencing approaches in plant species. In addition, the transcriptome reference was essential for gene model identification and quality assessment in the parallel project responsible for sequencing and assembly of the entire genome. In this study, the transcriptomic data were also used to address questions surrounding lineage-specific Dicer-like proteins in conifers. These proteins play a role in the control of transposable element proliferation and the related genome expansion in conifers.

English walnut (Juglans regia L.) is an economically important crop with > 99% of US walnuts produced in California. Changes in climate and recent drought cycles have raised concerns regarding the future of nut production and responsible water use in California agriculture. Our study used an association genetics approach to characterize ecophysiological traits such as water use efficiency as estimated by carbon isotope discrimination (Δ13C), and photosynthetic capacity through foliar nitrogen composition, in important individuals of the Walnut Improvement Program, located at the University of California, Davis. Stable isotope and leaf measurements of 241 mature trees, representing 60 scion genotypes in established orchards were sampled in 2015 and 2016, followed by genotyping with the Walnut Axiom 700 k SNP Array. A mean Δ13C of 21.7‰ (σ: 0.9‰) was calculated for all individuals, as well as a mean nitrogen/leaf area (N/area) of 3.0 gN/m2 (σ: 0.5 gN/m2). A Bayesian analysis utilizing genomic relationships revealed rankings of the most water use efficient accessions as Solano (95-011-16), 67-013 (unreleased cross), and Tulare (67-011). 126,554 SNPs were used in a two-step association genetics approach identifying four loci associated with Δ13C after correction for multiple testing. Investigation of identified loci revealed an annotation on the J. regia genome of protein FAR1-related sequence 5-like, related to abiotic stress response. For uncharacterized markers, homologs were identified in Arabidopsis for two loci, similarly related to drought stress. [ABSTRACT FROM AUTHOR]