Analysis of >3400 worldwide eggplant accessions reveals two independent domestication events and multiple migration-diversification routes.

Eggplant (Solanum melongena) is an important Solanaceous crop, widely cultivated and consumed in Asia, the Mediterranean basin, and Southeast Europe. Its domestication centers and migration and diversification routes are still a matter of debate. We report the largest georeferenced and genotyped collection to this date for eggplant and its wild relatives, consisting of 3499 accessions from seven worldwide genebanks, originating from 105 countries in five continents. The combination of genotypic and passport data points to the existence of at least two main centers of domestication, in Southeast Asia and the Indian subcontinent, with limited genetic exchange between them. The wild and weedy eggplant ancestor S. insanum shows admixture with domesticated S. melongena, similar to what was described for other fruit-bearing Solanaceous crops such as tomato and pepper and their wild ancestors. After domestication, migration and admixture of eggplant populations from different regions have been less conspicuous with respect to tomato and pepper, thus better preserving 'local' phenotypic characteristics. The data allowed the identification of misclassified and putatively duplicated accessions, facilitating genebank management. All the genetic, phenotypic, and passport data have been deposited in the Open Access G2P-SOL database, and constitute an invaluable resource for understanding the domestication, migration and diversification of this cosmopolitan vegetable.

Genetics and breeding of phenolic content in tomato, eggplant and pepper fruits.

Phenolic acids and flavonoids are large groups of secondary metabolites ubiquitous in the plant kingdom. They are currently in the spotlight due to the numerous health benefits associated with their consumption, as well as for their vital roles in plant biological processes and in plant-environment interaction. Tomato, eggplant and pepper are in the top ten most consumed vegetables in the world, and their fruit accumulation profiles have been extensively characterized, showing substantial differences. A broad array of genetic and genomic tools has helped to identify QTLs and candidate genes associated with the fruit biosynthesis of phenolic acids and flavonoids. The aim of this review was to synthesize the available information making it easily available for researchers and breeders. The phenylpropanoid pathway is tightly regulated by structural genes, which are conserved across species, along with a complex network of regulatory elements like transcription factors, especially of MYB family, and cellular transporters. Moreover, phenolic compounds accumulate in tissue-specific and developmental-dependent ways, as different paths of the metabolic pathway are activated/deactivated along with fruit development. We retrieved 104 annotated putative orthologues encoding for key enzymes of the phenylpropanoid pathway in tomato (37), eggplant (29) and pepper (38) and compiled 267 QTLs (217 for tomato, 16 for eggplant and 34 for pepper) linked to fruit phenolic acids, flavonoids and total phenolics content. Combining molecular tools and genetic variability, through both conventional and genetic engineering strategies, is a feasible approach to improve phenolics content in tomato, eggplant and pepper. Finally, although the phenylpropanoid biosynthetic pathway has been well-studied in the Solanaceae, more research is needed on the identification of the candidate genes behind many QTLs, as well as their interactions with other QTLs and genes.

CaMYB12-like underlies a major QTL for flavonoid content in pepper (Capsicum annuum) fruit.

The regulation of flavonoid biosynthesis is only partially explored in pepper (Capsicum annuum L.). The genetic basis underlying flavonoid variation in pepper fruit was studied. Variation of flavonoids in fruit of a segregating F2 population was studied using LC-MS followed by quantitative trait locus (QTL) analysis. Near-isogenic lines (NILs), BC1 S1 populations, virus-induced gene silenced (VIGS) and transgenic overexpression were used to confirm the QTL and the underlying candidate gene. A major QTL for flavonoid content was found in chromosome 5, and a CaMYB12-like transcription factor gene was identified as candidate gene. Near-isogenic lines (NILs) contrasting for CaMYB12-like confirmed its association with the flavonoid content variation. Virus-induced gene silencing (VIGS) of CaMYB12-like led to a significant decrease in the expression of several flavonoid pathway genes and a drastic decrease in flavonoid levels in silenced fruits. Expression of CaMYB12-like in the tomato slmyb12 mutant led to enhanced levels of several flavonoids in the fruit skin. Introgression of the CaMYB12-like allele into two cultivated varieties also increased flavonoid content in their fruits. A combination of metabolomic, genetic and gene functional analyses led to discovery of CaMYB12-like as a major regulator of flavonoid variation in pepper fruit and demonstrated its potential to breed for high-flavonoid content in cultivated pepper.

Identification of candidate genes associated with less-photosensitive anthocyanin phenotype using an EMS mutant (pind) in eggplant (Solanum melongena L.).

Eggplant (Solanum melongena L.) is a highly nutritious and economically important vegetable crop. However, the fruit peel of eggplant often shows poor coloration owing to low-light intensity during cultivation, especially in the winter. The less-photosensitive varieties produce anthocyanin in low light or even dark conditions, making them valuable breeding materials. Nevertheless, genes responsible for anthocyanin biosynthesis in less-photosensitive eggplant varieties are not characterized. In this study, an EMS mutant, named purple in the dark (pind), was used to identify the key genes responsible for less-photosensitive coloration. Under natural conditions, the peel color and anthocyanin content in pind fruits were similar to that of wildtype '14-345'. The bagged pind fruits were light purple, whereas those of '14-345' were white; and the anthocyanin content in the pind fruit peel was significantly higher than that in '14-345'. Genetic analysis revealed that the less-photosensitive trait was controlled by a single dominant gene. The candidate gene was mapped on chromosome 10 in the region 7.72 Mb to 11.71 Mb. Thirty-five differentially expressed genes, including 12 structural genes, such as CHS, CHI, F3H, DFR, ANS, and UFGT, and three transcription factors MYB113, GL3, and TTG2, were identified in pind using RNA-seq. Four candidate genes EGP21875 (myb domain protein 113), EGP21950 (unknown protein), EGP21953 (CAAX amino-terminal protease family protein), and EGP21961 (CAAX amino-terminal protease family protein) were identified as putative genes associated with less-photosensitive anthocyanin biosynthesis in pind. These findings may clarify the molecular mechanisms underlying less-photosensitive anthocyanin biosynthesis in eggplant.

Discovery of a DFR gene that controls anthocyanin accumulation in the spiny Solanum group: roles of a natural promoter variant and alternative splicing.

Anthocyanins are important pigments that impart color in plants. In Solanum, different species display various fruit or flower colors due to varying degrees of anthocyanin accumulation. Here we identified two anthocyanin-free mutants from an ethylmethane sulfonate-induced mutant library and naturally occurring mutants in Solanum melongena, with mutations in the 5' splicing site of the second intron of dihydroflavonol-4-reductase (DFR) - leading to altered splicing. Further study revealed that alternative splicing of the second intron was closely related to anthocyanin accumulation in 17 accessions from three cultivated species: S. melongena, Solanum macrocarpon and Solanum aethiopicum, and their wild related species. Analysis of natural variations of DFR, using an expanded population including 282 accessions belonging to the spiny Solanum group, identified a single-nucleotide polymorphism in the MYB recognition site in the promoter region, which causes differential expression of DFR and affects anthocyanin accumulation in fruits of the detected accessions. Our study suggests that, owing to years of domestication, the natural variation in the DFR promoter region and the alternative splicing of the DFR gene account for altered anthocyanin accumulation during spiny Solanum domestication.

Global range expansion history of pepper (Capsicum spp.) revealed by over 10,000 genebank accessions.

Genebanks collect and preserve vast collections of plants and detailed passport information, with the aim of preserving genetic diversity for conservation and breeding. Genetic characterization of such collections has the potential to elucidate the genetic histories of important crops, use marker-trait associations to identify loci controlling traits of interest, search for loci undergoing selection, and contribute to genebank management by identifying taxonomic misassignments and duplicates. We conducted a genomic scan with genotyping by sequencing (GBS) derived single nucleotide polymorphisms (SNPs) of 10,038 pepper (Capsicum spp.) accessions from worldwide genebanks and investigated the recent history of this iconic staple. Genomic data detected up to 1,618 duplicate accessions within and between genebanks and showed that taxonomic ambiguity and misclassification often involve interspecific hybrids that are difficult to classify morphologically. We deeply interrogated the genetic diversity of the commonly consumed Capsicum annuum to investigate its history, finding that the kinds of peppers collected in broad regions across the globe overlap considerably. The method ReMIXTURE-using genetic data to quantify the similarity between the complement of peppers from a focal region and those from other regions-was developed to supplement traditional population genetic analyses. The results reflect a vision of pepper as a highly desirable and tradable cultural commodity, spreading rapidly throughout the globe along major maritime and terrestrial trade routes. Marker associations and possible selective sweeps affecting traits such as pungency were observed, and these traits were shown to be distributed nonuniformly across the globe, suggesting that human preferences exerted a primary influence over domesticated pepper genetic structure.

The Genetic Basis of Tomato Aroma.

Tomato (Solanum lycopersicum L.) aroma is determined by the interaction of volatile compounds (VOCs) released by the tomato fruits with receptors in the nose, leading to a sensorial impression, such as "sweet", "smoky", or "fruity" aroma. Of the more than 400 VOCs released by tomato fruits, 21 have been reported as main contributors to the perceived tomato aroma. These VOCs can be grouped in five clusters, according to their biosynthetic origins. In the last decades, a vast array of scientific studies has investigated the genetic component of tomato aroma in modern tomato cultivars and their relatives. In this paper we aim to collect, compare, integrate and summarize the available literature on flavour-related QTLs in tomato. Three hundred and 5ifty nine (359) QTLs associated with tomato fruit VOCs were physically mapped on the genome and investigated for the presence of potential candidate genes. This review makes it possible to (i) pinpoint potential donors described in literature for specific traits, (ii) highlight important QTL regions by combining information from different populations, and (iii) pinpoint potential candidate genes. This overview aims to be a valuable resource for researchers aiming to elucidate the genetics underlying tomato flavour and for breeders who aim to improve tomato aroma.

Exploration of a Resequenced Tomato Core Collection for Phenotypic and Genotypic Variation in Plant Growth and Fruit Quality Traits.

A tomato core collection consisting of 122 gene bank accessions, including landraces, old cultivars, and wild relatives, was explored for variation in several plant growth, yield and fruit quality traits. The resequenced accessions were also genotyped with respect to a number of mutations or variations in key genes known to underlie these traits. The yield-related traits fruit number and fruit weight were much higher in cultivated varieties when compared to wild accessions, while, in wild tomato accessions, Brix was higher than in cultivated varieties. Known mutations in fruit size and shape genes could well explain the fruit size variation, and fruit colour variation could be well explained by known mutations in key genes of the carotenoid and flavonoid pathway. The presence and phenotype of several plant architecture affecting mutations, such as self-pruning (sp), compound inflorescence (s), jointless-2 (j-2), and potato leaf (c) were also confirmed. This study provides valuable phenotypic information on important plant growth- and quality-related traits in this collection. The allelic distribution of known genes that underlie these traits provides insight into the role and importance of these genes in tomato domestication and breeding. This resource can be used to support (precision) breeding strategies for tomato crop improvement.

Analysis of wild tomato introgression lines elucidates the genetic basis of transcriptome and metabolome variation underlying fruit traits and pathogen response.

Wild tomato species represent a rich gene pool for numerous desirable traits lost during domestication. Here, we exploited an introgression population representing wild desert-adapted species and a domesticated cultivar to establish the genetic basis of gene expression and chemical variation accompanying the transfer of wild-species-associated fruit traits. Transcriptome and metabolome analysis of 580 lines coupled to pathogen sensitivity assays resulted in the identification of genomic loci associated with levels of hundreds of transcripts and metabolites. These associations occurred in hotspots representing coordinated perturbation of metabolic pathways and ripening-related processes. Here, we identify components of the Solanum alkaloid pathway, as well as genes and metabolites involved in pathogen defense and linking fungal resistance with changes in the fruit ripening regulatory network. Our results outline a framework for understanding metabolism and pathogen resistance during tomato fruit ripening and provide insights into key fruit quality traits.

The genetic and functional analysis of flavor in commercial tomato: the FLORAL4 gene underlies a QTL for floral aroma volatiles in tomato fruit.

Tomato (Solanum lycopersicum L.) has become a popular model for genetic studies of fruit flavor in the last two decades. In this article we present a study of tomato fruit flavor, including an analysis of the genetic, metabolic and sensorial variation of a collection of contemporary commercial glasshouse tomato cultivars, followed by a validation of the associations found by quantitative trait locus (QTL) analysis of representative biparental segregating populations. This led to the identification of the major sensorial and chemical components determining fruit flavor variation and detection of the underlying QTLs. The high representation of QTL haplotypes in the breeders' germplasm suggests that there is great potential for applying these QTLs in current breeding programs aimed at improving tomato flavor. A QTL on chromosome 4 was found to affect the levels of the phenylalanine-derived volatiles (PHEVs) 2-phenylethanol, phenylacetaldehyde and 1-nitro-2-phenylethane. Fruits of near-isogenic lines contrasting for this locus and in the composition of PHEVs significantly differed in the perception of fruity and rose-hip-like aroma. The PHEV locus was fine mapped, which allowed for the identification of FLORAL4 as a candidate gene for PHEV regulation. Using a gene-editing-based (CRISPR-CAS9) reverse-genetics approach, FLORAL4 was demonstrated to be the key factor in this QTL affecting PHEV accumulation in tomato fruit.

The rin, nor and Cnr spontaneous mutations inhibit tomato fruit ripening in additive and epistatic manners.

Tomato fruit ripening is regulated by transcription factors (TFs), their downstream effector genes, and the ethylene biosynthesis and signalling pathway. Spontaneous non-ripening mutants ripening inhibitor (rin), non-ripening (nor) and Colorless non-ripening (Cnr) correspond with mutations in or near the TF-encoding genes MADS-RIN, NAC-NOR and SPL-CNR, respectively. Here, we produced heterozygous single and double mutants of rin, nor and Cnr and evaluated their functions and genetic interactions in the same genetic background. We showed how these mutations interact at the level of phenotype, individual effector gene expression, and sensory and quality aspects, in a dose-dependent manner. Rin and nor have broadly similar quantitative effects on all aspects, demonstrating their additivity in fruit ripening regulation. We also found that the Cnr allele is epistatic to rin and nor and that its pleiotropic effects on fruit size and volatile production, in contrast to the well-known dominant effect on ripening, are incompletely dominant, or recessive.

Reprogramming of Tomato Leaf Metabolome by the Activity of Heat Stress Transcription Factor HsfB1.

Plants respond to high temperatures with global changes of the transcriptome, proteome, and metabolome. Heat stress transcription factors (Hsfs) are the core regulators of transcriptome responses as they control the reprogramming of expression of hundreds of genes. The thermotolerance-related function of Hsfs is mainly based on the regulation of many heat shock proteins (HSPs). Instead, the Hsf-dependent reprogramming of metabolic pathways and their contribution to thermotolerance are not well described. In tomato (Solanum lycopersicum), manipulation of HsfB1, either by suppression or overexpression (OE) leads to enhanced thermotolerance and coincides with distinct profile of metabolic routes based on a metabolome profiling of wild-type (WT) and HsfB1 transgenic plants. Leaves of HsfB1 knock-down plants show an accumulation of metabolites with a positive effect on thermotolerance such as the sugars sucrose and glucose and the polyamine putrescine. OE of HsfB1 leads to the accumulation of products of the phenylpropanoid and flavonoid pathways, including several caffeoyl quinic acid isomers. The latter is due to the enhanced transcription of genes coding key enzymes in both pathways, in some cases in both non-stressed and stressed plants. Our results show that beyond the control of the expression of Hsfs and HSPs, HsfB1 has a wider activity range by regulating important metabolic pathways providing an important link between stress response and physiological tomato development.

Natural variation in HsfA2 pre-mRNA splicing is associated with changes in thermotolerance during tomato domestication.

Wild relatives of crops thrive in habitats where environmental conditions can be restrictive for productivity and survival of cultivated species. The genetic basis of this variability, particularly for tolerance to high temperatures, is not well understood. We examined the capacity of wild and cultivated accessions to acclimate to rapid temperature elevations that cause heat stress (HS). We investigated genotypic variation in thermotolerance of seedlings of wild and cultivated accessions. The contribution of polymorphisms associated with thermotolerance variation was examined regarding alterations in function of the identified gene. We show that tomato germplasm underwent a progressive loss of acclimation to strong temperature elevations. Sensitivity is associated with intronic polymorphisms in the HS transcription factor HsfA2 which affect the splicing efficiency of its pre-mRNA. Intron splicing in wild species results in increased synthesis of isoform HsfA2-II, implicated in the early stress response, at the expense of HsfA2-I which is involved in establishing short-term acclimation and thermotolerance. We propose that the selection for modern HsfA2 haplotypes reduced the ability of cultivated tomatoes to rapidly acclimate to temperature elevations, but enhanced their short-term acclimation capacity. Hence, we provide evidence that alternative splicing has a central role in the definition of plant fitness plasticity to stressful conditions.

The effect of isolation methods of tomato pollen on the results of metabolic profiling.

INTRODUCTION: Untargeted metabolomics is a powerful tool to detect hundreds of metabolites within a given tissue and to compare the metabolite composition of samples in a comprehensive manner. However, with regard to pollen research such comprehensive metabolomics approaches are yet not well developed. To enable isolation of pollen that is tightly enclosed within the anthers of the flower, such as immature pollen, the current pollen isolation protocols require the use of a watery solution. These protocols raise a number of concerns for their suitability in metabolomics analyses, in view of possible metabolic activities in the pollen and contamination with anther metabolites. OBJECTIVES: We assessed the effect of different sample preparation procedures currently used for pollen isolation for their suitability to perform metabolomics of tomato pollen. METHODS: Pollen were isolated using different methods and the metabolic profiles were analysed by liquid chromatography-mass spectrometry (LC-MS). RESULTS: Our results demonstrated that pollen isolation in a watery solution led to (i) rehydration of the pollen grains, inducing marked metabolic changes in flavonoids, phenylpropanoids and amino acids and thus resulting in a metabolite profile that did not reflect the one of mature dry pollen, (ii) hydrolysis of sucrose into glucose and fructose during subsequent metabolite extraction, unless the isolated and rehydrated pollen were lyophilized prior to extraction, and (iii) contamination with anther-specific metabolites, such as alkaloids, thus compromising the metabolic purity of the pollen fraction. CONCLUSION: We conclude that the current practices used to isolate pollen are suboptimal for metabolomics analyses and provide recommendations on how to improve the pollen isolation protocol, in order to obtain the most reliable metabolic profile from pollen tissue.

Breeding Has Increased the Diversity of Cultivated Tomato in The Netherlands.

It is generally believed that domestication and breeding of plants has led to genetic erosion, including loss of nutritional value and resistances to diseases, especially in tomato. We studied the diversity dynamics of greenhouse tomato varieties in NW Europe, especially The Netherlands, over the last seven decades. According to the used SNP array, the genetic diversity was indeed very low during the 1960s, but is now eight times higher when compared to that dip. The pressure since the 1970s to apply less pesticides led to the introgression of many disease resistances from wild relatives, representing the first boost of genetic diversity. In Europe a second boost ensued, largely driven by German popular media who named poor tasting tomatoes Wasserbomben (water bombs). The subsequent collapse of Dutch tomato exports to Germany fueled breeding for fruit flavor, further increasing diversity since the 1990s. The increased diversity in composition of aroma volatiles observed starting from 1990s may reflect the efforts of breeders to improve fruit quality. Specific groups of aroma compounds showed different quantitative trend over the decades studied. Our study provides compelling evidence that breeding has increased the diversity of tomato varieties considerably since the 1970s.

Anthocyanin Biosynthesis and Degradation Mechanisms in Solanaceous Vegetables: A Review.

Anthocyanins are a group of polyphenolic pigments that are ubiquitously found in the plant kingdom. In plants, anthocyanins play a role not only in reproduction, by attracting pollinators and seed dispersers, but also in protection against various abiotic and biotic stresses. There is accumulating evidence that anthocyanins have health-promoting properties, which makes anthocyanin metabolism an interesting target for breeders and researchers. In this review, the state of the art knowledge concerning anthocyanins in the Solanaceous vegetables, i.e., pepper, tomato, eggplant, and potato, is discussed, including biochemistry and biological function of anthocyanins, as well as their genetic and environmental regulation. Anthocyanin accumulation is determined by the balance between biosynthesis and degradation. Although the anthocyanin biosynthetic pathway has been well-studied in Solanaceous vegetables, more research is needed on the inhibition of biosynthesis and, in particular, the anthocyanin degradation mechanisms if we want to control anthocyanin content of Solanaceous vegetables. In addition, anthocyanin metabolism is distinctly affected by environmental conditions, but the molecular regulation of these effects is poorly understood. Existing knowledge is summarized and current gaps in our understanding are highlighted and discussed, to create opportunities for the development of anthocyanin-rich crops through breeding and environmental management.

Untargeted metabolomic analysis of tomato pollen development and heat stress response.

KEY MESSAGE: Pollen development metabolomics. Developing pollen is among the plant structures most sensitive to high temperatures, and a decrease in pollen viability is often associated with an alteration of metabolite content. Most of the metabolic studies of pollen have focused on a specific group of compounds, which limits the identification of physiologically important metabolites. To get a better insight into pollen development and the pollen heat stress response, we used a liquid chromatography-mass spectrometry platform to detect secondary metabolites in pollen of tomato (Solanum lycopersicum L.) at three developmental stages under control conditions and after a short heat stress at 38 °C. Under control conditions, the young microspores accumulated a large amount of alkaloids and polyamines, whereas the mature pollen strongly accumulated flavonoids. The heat stress treatment led to accumulation of flavonoids in the microspore. The biological role of the detected metabolites is discussed. This study provides the first untargeted metabolomic analysis of developing pollen under a changing environment that can serve as reference for further studies.

Identification of Loci Affecting Accumulation of Secondary Metabolites in Tomato Fruit of a Solanum lycopersicum × Solanum chmielewskii Introgression Line Population.

Semi-polar metabolites such as flavonoids, phenolic acids, and alkaloids are very important health-related compounds in tomato. As a first step to identify genes responsible for the synthesis of semi-polar metabolites, quantitative trait loci (QTLs) that influence the semi-polar metabolite content in red-ripe tomato fruit were identified, by characterizing fruits of a population of introgression lines (ILs) derived from a cross between the cultivated tomato Solanum lycopersicum and the wild species Solanum chmielewskii. By analyzing fruits of plants grown at two different locations, we were able to identify robust metabolite QTLs for changes in phenylpropanoid glycoconjugation on chromosome 9, for accumulation of flavonol glycosides on chromosome 5, and for alkaloids on chromosome 7. To further characterize the QTLs we used a combination of genome sequencing, transcriptomics and targeted metabolomics to identify candidate key genes underlying the observed metabolic variation.

HsfA2 Controls the Activity of Developmentally and Stress-Regulated Heat Stress Protection Mechanisms in Tomato Male Reproductive Tissues.

Male reproductive tissues are more sensitive to heat stress (HS) compared to vegetative tissues, but the basis of this phenomenon is poorly understood. Heat stress transcription factors (Hsfs) regulate the transcriptional changes required for protection from HS In tomato (Solanum lycopersicum), HsfA2 acts as coactivator of HsfA1a and is one of the major Hsfs accumulating in response to elevated temperatures. The contribution of HsfA2 in heat stress response (HSR) and thermotolerance was investigated in different tissues of transgenic tomato plants with suppressed HsfA2 levels (A2AS). Global transcriptome analysis and immunodetection of two major Hsps in vegetative and reproductive tissues showed that HsfA2 regulates subsets of HS-induced genes in a tissue-specific manner. Accumulation of HsfA2 by a moderate HS treatment enhances the capacity of seedlings to cope with a subsequent severe HS, suggesting an important role for HsfA2 in regulating acquired thermotolerance. In pollen, HsfA2 is an important coactivator of HsfA1a during HSR HsfA2 suppression reduces the viability and germination rate of pollen that received the stress during the stages of meiosis and microspore formation but had no effect on more advanced stages. In general, pollen meiocytes and microspores are characterized by increased susceptibility to HS due to their lower capacity to induce a strong HSR This sensitivity is partially mitigated by the developmentally regulated expression of HsfA2 and several HS-responsive genes mediated by HsfA1a under nonstress conditions. Thereby, HsfA2 is an important factor for the priming process that sustains pollen thermotolerance during microsporogenesis.

Genotype-phenotype modeling considering intermediate level of biological variation: a case study involving sensory traits, metabolites and QTLs in ripe tomatoes.

Modeling genotype-phenotype relationships is a central objective in plant genetics and breeding. Commonly, variations in phenotypic traits are modeled directly in relation to variations at the DNA level, regardless of intermediate levels of biological variation. Here we present an integrative method for the simultaneous modeling of a set of multilevel phenotypic responses to variations at the DNA level. More specifically, for ripe tomato fruits, we use Gaussian graphical models and causal inference techniques to learn the dependencies of 24 sensory traits on 29 metabolites and the dependencies of those sensory and metabolic traits on 21 QTLs. The inferred dependency network which, though not essentially representing biological pathways, suggests how the effects of allele substitutions propagate through multilevel phenotypes. Such simultaneous study of the underlying genetic architecture and multifactorial interactions is expected to enhance the prediction and manipulation of complex traits.