Insights into the early transcriptomic response against watermelon mosaic virus in melon.

BACKGROUND: Watermelon mosaic virus (WMV) is one of the most prevalent viruses affecting melon worldwide. Recessive resistance to WMV in melon has previously been reported in the African accession TGR-1551. Moreover, the genomic regions associated to the resistance have also been described. Nevertheless, the transcriptomic response that might infer the resistance to this potyvirus has not been explored. RESULTS: We have performed a comparative transcriptomic analysis using mock and WMV-inoculated plants of the susceptible cultivar "Bola de oro" (BO) and a resistant RIL (Recombinant inbred line) derived from the initial cross between "TGR-1551" and BO. In total, 616 genes were identified as differentially expressed and the weighted gene co-expression network analysis (WGCNA) detected 19 gene clusters (GCs), of which 7 were differentially expressed for the genotype x treatment interaction term. SNPs with a predicted high impact on the protein function were detected within the coding regions of most of the detected DEGs. Moreover, 3 and 16 DEGs were detected within the QTL regions previously described in chromosomes 11 and 5, respectively. In addition to these two specific genomic regions, we also observde large transcriptomic changes from genes spread across the genome in the resistant plants in response to the virus infection. This early response against WMV implied genes involved in plant-pathogen interaction, plant hormone signal transduction, the MAPK signaling pathway or ubiquitin mediated proteolysis, in detriment to the photosynthetic and basal metabolites pathways. Moreover, the gene MELO3C021395, which coded a mediator of RNA polymerase II transcription subunit 33A (MED33A), has been proposed as the candidate gene located on chromosome 11 conferring resistance to WMV. CONCLUSIONS: The comparative transcriptomic analysis presented here showed that, even though the resistance to WMV in TGR-1551 has a recessive nature, it triggers an active defense response at a transcriptomic level, which involves broad-spectrum resistance mechanisms. Thus, this study represents a step forward on our understanding of the mechanisms underlaying WMV resistance in melon. In addition, it sheds light into a broader topic on the mechanisms of recessive resistances.

Synthetic conversion of leaf chloroplasts into carotenoid-rich plastids reveals mechanistic basis of natural chromoplast development.

Plastids, the defining organelles of plant cells, undergo physiological and morphological changes to fulfill distinct biological functions. In particular, the differentiation of chloroplasts into chromoplasts results in an enhanced storage capacity for carotenoids with industrial and nutritional value such as beta-carotene (provitamin A). Here, we show that synthetically inducing a burst in the production of phytoene, the first committed intermediate of the carotenoid pathway, elicits an artificial chloroplast-to-chromoplast differentiation in leaves. Phytoene overproduction initially interferes with photosynthesis, acting as a metabolic threshold switch mechanism that weakens chloroplast identity. In a second stage, phytoene conversion into downstream carotenoids is required for the differentiation of chromoplasts, a process that involves a concurrent reprogramming of nuclear gene expression and plastid morphology for improved carotenoid storage. We hence demonstrate that loss of photosynthetic competence and enhanced production of carotenoids are not just consequences but requirements for chloroplasts to differentiate into chromoplasts.

A Novel Introgression Line Library Derived from a Wild Melon Gives Insights into the Genetics of Melon Domestication, Uncovering New Genetic Variability Useful for Breeding.

A collection of 30 melon introgression lines (ILs) was developed from the wild accession Ames 24297 (TRI) into 'Piel de Sapo' (PS) genetic background. Each IL carried an average of 1.4 introgressions from TRI, and the introgressions represented 91.4% of the TRI genome. Twenty-two ILs, representing 75% of the TRI genome, were evaluated in greenhouse (Algarrobo and Meliana) and field (Alcàsser) trials, mainly to study traits related to domestication syndrome such as fruit weight (FW) and flesh content (FFP), as well as other fruit quality traits as fruit shape (FS), flesh firmness (FF), soluble solid concentration (SSC), rind color and abscission layer. The IL collection showed an impressive variation in size-related traits, with FW ranging from 800 to 4100 g, reflecting the strong effect of the wild genome on these traits. Most of the ILs produced smaller fruits compared with PS; however, unexpectedly, the IL TRI05-2 produced bigger fruits, likely due to new epistatic interacions with the PS genetic background. In contrast, the genotypic effect for FS was smaller, and few QTLs with notable effects were detected. Interestingly, variability was also observed for FFP, FF and SSC, rind color and abscission layer formation. Genes in these introgressions are candidates for having been involved in melon domestication and diversification as well. These results confirm that the TRI IL collection is a very powerful tool for mapping traits of agronomic interest in melon, allowing the confirmation of previously reported QTLs and the identification of new ones to better understand the domestication process of this crop.

A cryptic variation in a member of the Ovate Family Proteins is underlying the melon fruit shape QTL fsqs8.1.

KEY MESSAGE: The gene underlying the melon fruit shape QTL fsqs8.1 is a member of the Ovate Family Proteins. Variation in fruit morphology is caused by changes in gene expression likely due to a cryptic structural variation in this locus. Melon cultivars have a wide range of fruit morphologies. Quantitative trait loci (QTL) have been identified underlying such diversity. This research focuses on the fruit shape QTL fsqs8.1, previously detected in a cross between the accession PI 124112 (CALC, producing elongated fruit) and the cultivar 'Piel de Sapo' (PS, producing oval fruit). The CALC fsqs8.1 allele induced round fruit shape, being responsible for the transgressive segregation for this trait observed in that population. In fact, the introgression line CALC8-1, carrying the fsqs8.1 locus from CALC into the PS genetic background, produced perfect round fruit. Following a map-based cloning approach, we found that the gene underlying fsqs8.1 is a member of the Ovate Family Proteins (OFP), CmOFP13, likely a homologue of AtOFP1 and SlOFP20 from Arabidopsis thaliana and tomato, respectively. The induction of the round shape was due to the higher expression of the CALC allele at the early ovary development stage. The fsqs8.1 locus showed an important structural variation, being CmOFP13 surrounded by two deletions in the CALC genome. The deletions are present at very low frequency in melon germplasm. Deletions and single nucleotide polymorphisms in the fsqs8.1 locus could not be not associated with variation in fruit shape among different melon accessions, what indicates that other genetic factors should be involved to induce the CALC fsqs8.1 allele effects. Therefore, fsqs8.1 is an example of a cryptic variation that alters gene expression, likely due to structural variation, resulting in phenotypic changes in melon fruit morphology.

Advanced Genetic Studies on Powdery Mildew Resistance in TGR-1551.

Cucurbits powdery mildew (CPM) is one of the main limiting factors of melon cultivation worldwide. Resistance to races 1, 2, and 5 has been reported in the African accession TGR-1551, whose resistance is controlled by a dominant-recessive epistasis. The dominant and recessive quantitative trail loci (QTL) have previously been located in chromosomes 5 and 12, respectively. We used several densely genotyped BC3 families derived from the cross between TGR-1551 and the susceptible cultivar 'Bola de Oro' to finely map these resistance regions. The further phenotyping and genotyping of the selected BC5, BC5S1, BC5S2, BC4S1, BC4xPS, and (BC4xPS) S1 offspring allowed for the narrowing of the candidate intervals to a 250 and 381 kb region in chromosomes 5 and 12, respectively. Moreover, the temperature effect over the resistance provided by the dominant gene has been confirmed. High resolution melting markers (HRM) were tightly linked to both resistance regions and will be useful in marker-assisted selection programs. Candidate R genes with variants between parents that caused a potential modifier impact on the protein function were identified within both intervals. These candidate genes provide targets for future functional analyses to better understand the resistance to powdery mildew in melons.

Spanish Melon Landraces: Revealing Useful Diversity by Genomic, Morphological, and Metabolomic Analysis.

Spain is a secondary centre of the diversification of the melon (Cucumis melo L.), with high diversity represented in highly appreciated landraces belonging to the Flexuosus and Ibericus groups. A collection of 47 accessions of Flexuosus, Chate, Piel de Sapo, Tendral, Amarillo, Blanco, and Rochet was analysed using a genotyping-by-sequencing (GBS) approach. A total of 66,971 quality SNPs were identified. Genetic analysis differentiated Ibericus accessions and exotic materials (Ameri, Momordica, Kachri, and Agrestis), while Flexuous accessions shared ancestry between them. Within the Ibericus group, no clear genomic distinction could be identified for the different landraces evaluated, with accessions of different landraces showing high genetic similarity. The morphological characterization confirmed that the external colour and fruit shape had been used as recognition patterns for Spanish melon landraces, but variability within a landrace exists. Differences were found in the sugars and acid and volatile profiles of the materials. Flexuosus and Chate melons at the immature commercial stage accumulated malic acid and low levels of hexoses, while Ibericus melons accumulated high contents of sucrose and citric acid. Specific trends could be identified in the Ibericus landraces. Tendral accumulated low levels of sugars and citric acid and high of malic acid, maintaining higher firmness, Rochet reached higher levels of sugars, and Amarillo tended to lower malic acid contents. Interestingly, high variability was found within landraces for the acidic profile, offering possibilities to alter taste tinges. The main volatile organic compounds (VOCs) in Flexuosus and Chate were aldehydes and alcohols, with clear differences between both groups. In the Ibericus landraces, general trends for VOC accumulation could be identified, but, again, a high level of variation exists. This situation highlights the necessity to develop depuration programs to promote on-farm in situ conservation and, at the same time, offers opportunities to establish new breeding program targets and to take advantage of these sources of variation.

A comprehensive RNA-Seq-based gene expression atlas of the summer squash (Cucurbita pepo) provides insights into fruit morphology and ripening mechanisms.

BACKGROUND: Summer squash (Cucurbita pepo: Cucurbitaceae) are a popular horticultural crop for which there is insufficient genomic and transcriptomic information. Gene expression atlases are crucial for the identification of genes expressed in different tissues at various plant developmental stages. Here, we present the first comprehensive gene expression atlas for a summer squash cultivar, including transcripts obtained from seeds, shoots, leaf stem, young and developed leaves, male and female flowers, fruits of seven developmental stages, as well as primary and lateral roots. RESULTS: In total, 27,868 genes and 2352 novel transcripts were annotated from these 16 tissues, with over 18,000 genes common to all tissue groups. Of these, 3812 were identified as housekeeping genes, half of which assigned to known gene ontologies. Flowers, seeds, and young fruits had the largest number of specific genes, whilst intermediate-age fruits the fewest. There also were genes that were differentially expressed in the various tissues, the male flower being the tissue with the most differentially expressed genes in pair-wise comparisons with the remaining tissues, and the leaf stem the least. The largest expression change during fruit development was early on, from female flower to fruit two days after pollination. A weighted correlation network analysis performed on the global gene expression dataset assigned 25,413 genes to 24 coexpression groups, and some of these groups exhibited strong tissue specificity. CONCLUSIONS: These findings enrich our understanding about the transcriptomic events associated with summer squash development and ripening. This comprehensive gene expression atlas is expected not only to provide a global view of gene expression patterns in all major tissues in C. pepo but to also serve as a valuable resource for functional genomics and gene discovery in Cucurbitaceae.

Re-evaluation of the role of Indian germplasm as center of melon diversification based on genotyping-by-sequencing analysis.

BACKGROUND: The importance of Indian germplasm as origin and primary center of diversity of cultivated melon is widely accepted. Genetic diversity of several collections from Indian has been studied previously, although an integrated analysis of these collections in a global diversity perspective has not been possible. In this study, a sample of Indian collections together with a selection of world-wide cultivars to analyze the genetic diversity structure based on Genotype by Sequence data. RESULTS: A set of 6158 informative Single Nucleotide Polymorphism (SNP) in 175 melon accessions was generated. Melon germplasm could be classified into six major groups, in concordance with horticultural groups. Indian group was in the center of the diversity plot, with the highest genetic diversity. No strict genetic differentiation between wild and cultivated accessions was appreciated in this group. Genomic regions likely involved in the process of diversification were also found. Interestingly, some SNPs differentiating inodorus and cantalupensis groups are linked to Quantitiative Trait Loci involved in ripening behavior (a major characteristic that differentiate those groups). Linkage disequilibrium was found to be low (17 kb), with more rapid decay in euchromatic (8 kb) than heterochromatic (30 kb) regions, demonstrating that recombination events do occur within heterochromatn, although at lower frequency than in euchromatin. Concomitantly, haplotype blocks were relatively small (59 kb). Some of those haplotype blocks were found fixed in different melon groups, being therefore candidate regions that are involved in the diversification of melon cultivars. CONCLUSIONS: The results support the hypothesis that India is the primary center of diversity of melon, Occidental and Far-East cultivars have been developed by divergent selection. Indian germplasm is genetically distinct from African germplasm, supporting independent domestication events. The current set of traditional Indian accessions may be considered as a population rather than a standard collection of fixed landraces with high intercrossing between cultivated and wild melons.

First RNA-seq approach to study fruit set and parthenocarpy in zucchini (Cucurbita pepo L.).

BACKGROUND: Zucchini fruit set can be limited due to unfavourable environmental conditions in off-seasons crops that caused ineffective pollination/fertilization. Parthenocarpy, the natural or artificial fruit development without fertilization, has been recognized as an important trait to avoid this problem, and is related to auxin signalling. Nevertheless, differences found in transcriptome analysis during early fruit development of zucchini suggest that other complementary pathways could regulate fruit formation in parthenocarpic cultivars of this species. The development of next-generation sequencing technologies (NGS) as RNA-sequencing (RNA-seq) opens a new horizon for mapping and quantifying transcriptome to understand the molecular basis of pathways that could regulate parthenocarpy in this species. The aim of the current study was to analyze fruit transcriptome of two cultivars of zucchini, a non-parthenocarpic cultivar and a parthenocarpic cultivar, in an attempt to identify key genes involved in parthenocarpy. RESULTS: RNA-seq analysis of six libraries (unpollinated, pollinated and auxin treated fruit in a non-parthenocarpic and parthenocarpic cultivar) was performed mapping to a new version of C. pepo transcriptome, with a mean of 92% success rate of mapping. In the non-parthenocarpic cultivar, 6479 and 2186 genes were differentially expressed (DEGs) in pollinated fruit and auxin treated fruit, respectively. In the parthenocarpic cultivar, 10,497 in pollinated fruit and 5718 in auxin treated fruit. A comparison between transcriptome of the unpollinated fruit for each cultivar has been performed determining that 6120 genes were differentially expressed. Annotation analysis of these DEGs revealed that cell cycle, regulation of transcription, carbohydrate metabolism and coordination between auxin, ethylene and gibberellin were enriched biological processes during pollinated and parthenocarpic fruit set. CONCLUSION: This analysis revealed the important role of hormones during fruit set, establishing the activating role of auxins and gibberellins against the inhibitory role of ethylene and different candidate genes that could be useful as markers for parthenocarpic selection in the current breeding programs of zucchini.

ETHQV6.3 is involved in melon climacteric fruit ripening and is encoded by a NAC domain transcription factor.

Fruit ripening is divided into climacteric and non-climacteric types depending on the presence or absence of a transient rise in respiration rate and the production of autocatalytic ethylene. Melon is ideal for the study of fruit ripening, as both climacteric and non-climacteric varieties exist. Two introgressions of the non-climacteric accession PI 161375, encompassed in the QTLs ETHQB3.5 and ETHQV6.3, into the non-climacteric 'Piel de Sapo' background are able to induce climacteric ripening independently. We report that the gene underlying ETHQV6.3 is MELO3C016540 (CmNAC-NOR), encoding a NAC (NAM, ATAF1,2, CUC2) transcription factor that is closely related to the tomato NOR (non-ripening) gene. CmNAC-NOR was functionally validated through the identification of two TILLING lines carrying non-synonymous mutations in the conserved NAC domain region. In an otherwise highly climacteric genetic background, both mutations provoked a significant delay in the onset of fruit ripening and in the biosynthesis of ethylene. The PI 161375 allele of ETHQV6.3 is similar to that of climacteric lines of the cantalupensis type and, when introgressed into the non-climacteric 'Piel de Sapo', partially restores its climacteric ripening capacity. CmNAC-NOR is expressed in fruit flesh of both climacteric and non-climacteric lines, suggesting that the causal mutation may not be acting at the transcriptional level. The use of a comparative genetic approach in a species with both climacteric and non-climacteric ripening is a powerful strategy to dissect the complex mechanisms regulating the onset of fruit ripening.

De novo assembly of the zucchini genome reveals a whole-genome duplication associated with the origin of the Cucurbita genus.

The Cucurbita genus (squashes, pumpkins and gourds) includes important domesticated species such as C. pepo, C. maxima and C. moschata. In this study, we present a high-quality draft of the zucchini (C. pepo) genome. The assembly has a size of 263 Mb, a scaffold N50 of 1.8 Mb and 34 240 gene models. It includes 92% of the conserved BUSCO core gene set, and it is estimated to cover 93.0% of the genome. The genome is organized in 20 pseudomolecules that represent 81.4% of the assembly, and it is integrated with a genetic map of 7718 SNPs. Despite the small genome size, three independent lines of evidence support that the C. pepo genome is the result of a whole-genome duplication: the topology of the gene family phylogenies, the karyotype organization and the distribution of 4DTv distances. Additionally, 40 transcriptomes of 12 species of the genus were assembled and analysed together with all the other published genomes of the Cucurbitaceae family. The duplication was detected in all the Cucurbita species analysed, including C. maxima and C. moschata, but not in the more distant cucurbits belonging to the Cucumis and Citrullus genera, and it is likely to have occurred 30 ± 4 Mya in the ancestral species that gave rise to the genus.

Repeated domestication of melon (Cucumis melo) in Africa and Asia and a new close relative from India.

PREMISE OF THE STUDY: The domestication history of melon is still unclear. An African or Asian origin has been suggested, but its closest wild relative was recently revealed to be an Australian species. The complicated taxonomic history of melon has resulted in additional confusion, with a high number of misidentified germplasm collections currently used by breeders and in genomics research. METHODS: Using seven DNA regions sequenced for 90% of the genus and the major cultivar groups, we sort out described names and infer evolutionary origins and domestication centers. KEY RESULTS: We found that modern melon cultivars go back to two lineages, which diverged ca. 2 million years ago. One is restricted to Asia (Cucumis melo subsp. melo), and the second, here described as C. melo subsp. meloides, is restricted to Africa. The Asian lineage has given rise to the widely commercialized cultivar groups and their market types, while the African lineage gave rise to cultivars still grown in the Sudanian region. We show that C. trigonus, an overlooked perennial and drought-tolerant species from India is among the closest living relatives of C. melo. CONCLUSIONS: Melon was domesticated at least twice: in Africa and Asia. The African lineage and the Indian C. trigonus are exciting new resources for breeding of melons tolerant to climate change.

An SNP-based saturated genetic map and QTL analysis of fruit-related traits in Zucchini using Genotyping-by-sequencing.

BACKGROUND: Cucurbita pepo is a cucurbit with growing economic importance worldwide. Zucchini morphotype is the most important within this highly variable species. Recently, transcriptome and Simple Sequence Repeat (SSR)- and Single Nucleotide Polymorphism (SNP)-based medium density maps have been reported, however further genomic tools are needed for efficient molecular breeding in the species. Our objective is to combine currently available complete transcriptomes and the Zucchini genome sequence with high throughput genotyping methods, mapping population development and extensive phenotyping to facilitate the advance of genomic research in this species. RESULTS: We report the Genotyping-by-sequencing analysis of a RIL population developed from the inter subspecific cross Zucchini x Scallop (ssp. pepo x ssp. ovifera). Several thousands of SNP markers were identified and genotyped, followed by the construction of a high-density linkage map based on 7,718 SNPs (average of 386 markers/linkage group) covering 2,817.6 cM of the whole genome, which is a great improvement with respect to previous maps. A QTL analysis was performed using phenotypic data obtained from the RIL population from three environments. In total, 48 consistent QTLs for vine, flowering and fruit quality traits were detected on the basis of a multiple-environment analysis, distributed in 33 independent positions in 15 LGs, and each QTL explained 1.5-62.9% of the phenotypic variance. Eight major QTLs, which could explain greater than 20% of the phenotypic variation were detected and the underlying candidate genes identified. CONCLUSIONS: Here we report the first SNP saturated map in the species, anchored to the physical map. Additionally, several consistent QTLs related to early flowering, fruit shape and length, and rind and flesh color are reported as well as candidate genes for them. This information will enhance molecular breeding in C. pepo and will assist the gene cloning underlying the studied QTLs, helping to reveal the genetic basis of the studied processes in squash.

Quantitative trait loci analysis of melon (Cucumis melo L.) domestication-related traits.

KEY MESSAGE: Loci on LGIV, VI, and VIII of melon genome are involved in the control of fruit domestication-related traits and they are candidate to have played a role in the domestication of the crop. The fruit of wild melons is very small (20-50 g) without edible pulp, contrasting with the large size and high pulp content of cultivated melon fruits. An analysis of quantitative trait loci (QTL) controlling fruit morphology domestication-related traits was carried out using an in vitro maintained F2 population from the cross between the Indian wild melon "Trigonus" and the western elite cultivar 'Piel de Sapo'. Twenty-seven QTL were identified in at least two out of the three field trials. Six of them were also being detected in BC1 and BC3 populations derived from the same cross. Ten of them were related to fruit morphological traits, 12 to fruit size characters, and 5 to pulp content. The Trigonus alleles decreased the value of the characters, except for the QTL at andromonoecious gene at linkage group (LG) II, and the QTL for pulp content at LGV. QTL genotypes accounted for a considerable degree of the total phenotypic variation, reaching up to 46%. Around 66% of the QTL showed additive gene action, 19% exhibited dominance, and 25% consisted of overdominance. The regions on LGIV, VI, and VIII included the QTL with more consistent and strong effects on domestication-related traits. QTLs on those regions were validated in BC2S1, BC2S2, and BC3 families, with "Trigonus" allele decreasing the fruit morphological traits in all cases. The validated QTL could represent loci involved in melon domestication, although further experiments as genomic variation studies across wild and cultivated genotypes would be necessary to confirm this hypothesis.

Resistance to tomato leaf curl New Delhi virus in melon is controlled by a major QTL located in chromosome 11.

KEY MESSAGE: Identification of three genomic regions and underlying candidate genes controlling the high level of resistance to ToLCNDV derived from a wild melon. SNP markers appropriated for MAS management of resistance. Tomato leaf curl New Delhi virus (ToLCNDV) is a bipartite begomovirus that severely affects melon crop (Cucumis melo) in the main production areas of Spain since 2012. In this work, we evaluated the degree of resistance of four accessions (two belonging to the subsp. agrestis var. momordica and two to the wild agrestis group) and their corresponding hybrids with a susceptible commercial melon belonging to the subsp. melo (Piel de Sapo, PS). The analysis using quantitative PCR (qPCR) allowed us to select one wild agrestis genotype (WM-7) with a high level of resistance and use it to construct segregating populations (F 2 and backcrosses). These populations were phenotyped for symptom severity and virus content using qPCR, and genotyped with different sets of SNP markers. Phenotyping and genotyping results in the F 2 and BC1s populations derived from the WM-7 × PS cross were used for QTL analysis. Three genomic regions controlling resistance to ToLCNDV were found, one major locus in chromosome 11 and two additional regions in chromosomes 12 and 2. The highest level of resistance (no or mild symptoms and very low viral titer) was obtained with the homozygous WM-7WM-7 genotype at the major QTL in chromosome 11, even with PSPS genotypes at the other two loci. The resistance derived from WM-7 is useful to develop new melon cultivars and the linked SNPs selected in this paper will be highly useful in marker-assisted breeding for ToLCNDV resistance in melon.

A new genomic library of melon introgression lines in a cantaloupe genetic background for dissecting desirable agronomical traits.

BACKGROUND: Genomic libraries of introgression lines (ILs) consist of collections of homozygous lines with a single chromosomal introgression from a donor genotype in a common, usually elite, genetic background, representing the whole donor genome in the full collection. Currently, the only available melon IL collection was generated using Piel de sapo (var. inodorus) as the recurrent background. ILs are not available in genetic backgrounds representing other important market class cultivars, such as the cantalupensis. The recent availability of genomic tools in melon, such as SNP collections and genetic maps, facilitates the development of such mapping populations. RESULTS: We have developed a new genomic library of introgression lines from the Japanese cv. Ginsen Makuwa (var. makuwa) into the French Charentais-type cv. Vedrantais (var. cantalupensis) genetic background. In order to speed up the breeding program, we applied medium-throughput SNP genotyping with Sequenom MassARRAY technology in early backcross generations and High Resolution Melting in the final steps. The phenotyping of the backcross generations and of the final set of 27 ILs (averaging 1.3 introgressions/plant and covering nearly 100 % of the donor genome), in three environments, allowed the detection of stable QTLs for flowering and fruit quality traits, including some that affect fruit size in chromosomes 6 and 11, others that change fruit shape in chromosomes 7 and 11, others that change flesh color in chromosomes 2, 8 and 9, and still others that increase sucrose content and delay climacteric behavior in chromosomes 5 and 10. CONCLUSIONS: A new melon IL collection in the Charentais genetic background has been developed. Genomic regions that consistently affect flowering and fruit quality traits have been identified, which demonstrates the suitability of this collection for dissecting complex traits in melon. Additionally, pre-breeding lines with new, commercially interesting phenotypes have been observed, including delayed climacteric ripening associated to higher sucrose levels, which is of great interest for Charentais cultivar breeding.

Variability of candidate genes, genetic structure and association with sugar accumulation and climacteric behavior in a broad germplasm collection of melon (Cucumis melo L.).

BACKGROUND: A collection of 175 melon (Cucumis melo L.) accessions (including wild relatives, feral types, landraces, breeding lines and commercial cultivars) from 50 countries was selected to study the phenotypic variability for ripening behavior and sugar accumulation. The variability of single nucleotide polymorphisms (SNPs) at 53 selected candidate genes involved in sugar accumulation and fruit ripening processes was studied, as well as their association with phenotypic variation of related traits. RESULTS: The collection showed a strong genetic structure, defining seven groups plus a number of accessions that could not be associated to any of the groups (admixture), which fitted well with the botanical classification of melon varieties. The variability in candidate genes for ethylene, cell wall and sugar-related traits was high and similar to SNPs located in reference genes. Variability at ripening candidate genes had an important weight on the genetic stratification of melon germplasm, indicating that traditional farmers might have selected for ripening traits during cultivar diversification. A strong relationship was also found between the genetic structure and phenotypic diversity, which could hamper genetic association studies. Accessions belonging to the ameri group are the most appropriate for association analysis given the high phenotypic and molecular diversity within the group, and lack of genetic structure. The most remarkable association was found between sugar content and SNPs in LG III, where a hotspot of sugar content QTLs has previously been defined. By studying the differences in allelic variation of SNPs within horticultural groups with specific phenotypic features, we also detected differential variation in sugar-related candidates located in LGIX and LGX, and in ripening-related candidates located in LGII and X, all in regions with previously mapped QTLs for the corresponding traits. CONCLUSIONS: In the current study we have found an important variability at both the phenotypic and candidate gene levels for ripening behavior and sugar accumulation in melon fruit. By combination of differences in allelic diversity and association analysis, we have identified several candidate genes that may be involved in the melon phenotypic diversity.

The genome of melon (Cucumis melo L.).

We report the genome sequence of melon, an important horticultural crop worldwide. We assembled 375 Mb of the double-haploid line DHL92, representing 83.3% of the estimated melon genome. We predicted 27,427 protein-coding genes, which we analyzed by reconstructing 22,218 phylogenetic trees, allowing mapping of the orthology and paralogy relationships of sequenced plant genomes. We observed the absence of recent whole-genome duplications in the melon lineage since the ancient eudicot triplication, and our data suggest that transposon amplification may in part explain the increased size of the melon genome compared with the close relative cucumber. A low number of nucleotide-binding site-leucine-rich repeat disease resistance genes were annotated, suggesting the existence of specific defense mechanisms in this species. The DHL92 genome was compared with that of its parental lines allowing the quantification of sequence variability in the species. The use of the genome sequence in future investigations will facilitate the understanding of evolution of cucurbits and the improvement of breeding strategies.

Involvement of ethylene biosynthesis and signalling in fruit set and early fruit development in zucchini squash (Cucurbita pepo L.).

BACKGROUND: We have identified a kind of parthenocarpy in zucchini squash which is associated with an incomplete andromonoecy, i.e. a partial conversion of female into bisexual flowers. Given that andromonoecy in this and other cucurbit species is caused by a reduction of ethylene production in the female flower, the associated parthenocarpic development of the fruit suggested the involvement of ethylene in fruit set and early fruit development. RESULTS: We have compared the production of ethylene as well as the expression of 13 ethylene biosynthesis and signalling genes in pollinated and unpollinated ovaries/fruits of two cultivars, one of which is parthenocarpic (Cavili), while the other is non-parthenocarpic (Tosca). In the latter, unpollinated ovaries show an induction of ethylene biosynthesis and ethylene signal transduction pathway genes three days after anthesis, which is concomitant with the initiation of fruit abortion and senescence. Fruit set and early fruit development in pollinated flowers of both cultivars and unpollinated flowers of Cavili is coupled with low ethylene biosynthesis and signalling, which would also explain the partial andromonoecy in the parthenocarpic genotype. The reduction of ethylene production in the ovary cosegregates with parthenocarpy and partial andromonoecy in the selfing progeny of Cavili. Moreover, the induction of ethylene in anthesis (by ethephon treatments) reduced the percentage of bisexual parthenocarpic flowers in Cavili, while the inhibition of ethylene biosynthesis or response (by AVG and STS treatments) induces not only andromonoecy but also the parthenocarpic development of the fruit in both cultivars. CONCLUSIONS: Results demonstrate that a reduction of ethylene production or signalling in the zucchini flower is able to induce fruit set and early fruit development, and therefore that ethylene is actively involved in fruit set and early fruit development. Auxin and TIBA treatments, inducing fruit set and early fruit development in this species, also inhibit ethylene production and the expression of ethylene biosynthesis and response genes. A model is presented that discusses the crosstalk between ethylene and auxin in the control of fruit set and early fruit development in zucchini squash.

SNP genotyping in melons: genetic variation, population structure, and linkage disequilibrium.

Novel sequencing technologies were recently used to generate sequences from multiple melon (Cucumis melo L.) genotypes, enabling the in silico identification of large single nucleotide polymorphism (SNP) collections. In order to optimize the use of these markers, SNP validation and large-scale genotyping are necessary. In this paper, we present the first validated design for a genotyping array with 768 SNPs that are evenly distributed throughout the melon genome. This customized Illumina GoldenGate assay was used to genotype a collection of 74 accessions, representing most of the botanical groups of the species. Of the assayed loci, 91 % were successfully genotyped. The array provided a large number of polymorphic SNPs within and across accessions. This set of SNPs detected high levels of variation in accessions from this crop's center of origin as well as from several other areas of melon diversification. Allele distribution throughout the genome revealed regions that distinguished between the two main groups of cultivated accessions (inodorus and cantalupensis). Population structure analysis showed a subdivision into five subpopulations, reflecting the history of the crop. A considerably low level of LD was detected, which decayed rapidly within a few kilobases. Our results show that the GoldenGate assay can be used successfully for high-throughput SNP genotyping in melon. Since many of the genotyped accessions are currently being used as the parents of breeding populations in various programs, this set of mapped markers could be used for future mapping and breeding efforts.