Abstract Heterosis, the superiority of hybrids over their parents, is a major genetic force associated with plant fitness and crop yield enhancement. We previously discovered and characterized root-mediated yield heterosis (RMYH) in melon ( Cucumis melo ) using a half-diallel population, derived from 20 diverse parents. In the current study we investigated the genetic architecture of RMYH using a segregating population derived from a selected F 1 hybrid (HDA019) that consistently induced RMYH under several melon scion varieties and growing conditions. 78 recombinant inbred lines (RILs) and their test-crosses to both parents were analyzed in yield trials as rootstocks under a common commercial scion variety. The population displayed normal root-mediated yield distribution and transgressive segregation relative to the parents but none of the RILs equaled the superior performance of the F 1 hybrid. RMYH of HDA019 was dissected to small effect QTLs showing mostly additive or dominant mode-of-inheritance and favorable QTL-alleles were contributed by both parents. Five consistent QTLs were selected and used to demonstrate the potential of root-mediated yield QTL pyramiding, and 20 combinations of QTL pairs and triplets supported the cumulative model for heterosis. Favorable QTLs alleles were introgressed to generate advanced QTL-backcross lines that were used for validation. This study provides first detailed genetic dissection of yield-related rootstock traits in cucurbits, highlighting rootstock breeding as an important underutilized route for improving yield and stress tolerance of crops. Key message Root–mediated yield heterosis in melon was genetically dissected using grafting strategy, revealing additive QTLs from both parents of the mapping population. Rootstock breeding through pyramiding of favorable alleles is proposed as strategy for enhancing crop yield and stress tolerance.

Through meta-genetic analysis of Cucumis melo sweetness, we expand the description of the complex genetic architecture of this trait. Integration of extensive new results with published QTL data provides an outline towards construction of a melon sweetness pan-QTLome. An ultimate objective in crop genetics is describing the complete repertoire of genes and alleles that shape the phenotypic variation of a quantitative trait within a species. Flesh sweetness is a primary determinant of fruit quality and consumer acceptance of melons. Cucumis melo is a diverse species that, among other traits, displays extensive variation in total soluble solids (TSS) content in fruit flesh, ranging from 20 Brix in non-sweet to 180 Brix in sweet accessions. We present here meta-genetic analysis of TSS and sugar variation in melon, using six different populations and fruit measurements collected from more than 30,000 open-field and greenhouse-grown plants, integrated with 15 published melon sweetness-related quantitative trait loci (QTL) studies. Starting with characterization of sugar composition variation across 180 diverse accessions that represent 3 subspecies and 12 of their cultivar-groups, we mapped TSS and sugar QTLs, and confirmed that sucrose accumulation is the key variable explaining TSS variation. All modes-of-inheritance for TSS were displayed by multi-season analysis of a broad half-diallel population derived from 20 diverse founders, with significant prevalence of the additive component. Through parallel genetic mapping in four advanced bi-parental populations, we identified common as well as unique TSS QTLs in 12 chromosomal regions. We demonstrate the cumulative less-than-additive nature of favorable TSS QTL alleles and the potential of a QTL-stacking approach. Using our broad dataset, we were additionally able to show that TSS variation displays weak genetic correlations with melon fruit size and ripening behavior, supporting effective breeding for sweetness per se. Our integrated analysis, combined with additional layers of published QTL data, broadens the perspective on the complex genetic landscape of melon sweetness and proposes a scheme towards future construction of a crop community-driven melon sweetness pan-QTLome.

Earliness and ripening behavior are important attributes of fruits on and off the vine, and affect quality and preference of both growers and consumers. Fruit ripening is a complex physiological process that involves metabolic shifts affecting fruit color, firmness, and aroma production. Melon is a promising model crop for the study of fruit ripening, as the full spectrum of climacteric behavior is represented across the natural variation. Using Recombinant Inbred Lines (RILs) population derived from the parental lines "Dulce" (reticulatus, climacteric) and "Tam Dew" (inodorus, non-climacteric) that vary in earliness and ripening traits, we mapped QTLs for ethylene emission, fruit firmness and days to flowering and maturity. To further annotate the main QTL intervals and identify candidate genes, we used Oxford Nanopore long-read sequencing in combination with Illumina short-read resequencing, to assemble the parental genomes de-novo. In addition to 2.5 million genome-wide SNPs and short InDels detected between the parents, we also highlight here the structural variation between these lines and the reference melon genome. Through systematic multi-layered prioritization process, we identified 18 potential polymorphisms in candidate genes within multi-trait QTLs. The associations of selected SNPs with earliness and ripening traits were further validated across a panel of 177 diverse melon accessions and across a diallel population of 190 F1 hybrids derived from a core subset of 20 diverse parents. The combination of advanced genomic tools with diverse germplasm and targeted mapping populations is demonstrated as a way to leverage forward genetics strategies to dissect complex horticulturally important traits.

Linking between genotype and phenotype is a fundamental goal in biology and requires robust data for both layers. The prominent increase in plant genome sequencing and comparisons of multiple related individuals, exposed the abundance of structural genomic variation and suggest that a single reference genome cannot represent the complete sequence diversity of a crop species, leading to the expansion of the pan-genome concept. For high-resolution forward genetics, this unprecedented access to genomic variation should be paralleled by availability and phenotypic characterization of genetic diversity, and effective integration between these layers. Here, we describe a multi-parental framework for trait dissection in melon, leveraging a novel pan-genome constructed for this crop. Melon (Cucumis melo L.) is an important crop from the Cucurbitaceae family, which display extensive phenotypic variation available for breeding. A diverse core set of 25 founder lines (MelonCore25) was sequenced using a combination of short and long-read technologies and their genomes were assembled de novo. The construction of a melon pan-genome exposed substantial variation in genome size and structure, including detection of ~300,000 structural variants and ~9 million SNPs. A half-diallel derived set of 300 F2 populations representing all possible MelonCore25 parental combinations was constructed as framework for trait dissection through integration with the pan-genome. We demonstrate the potential of this unified framework for genetic analysis of various melon traits, including rind color and mottling pattern, fruit sugar content and resistance to fungal diseases. We anticipate that utilization of this integrated resource will enhance genetic dissection of important traits and accelerate melon breeding. Significance statement Pan-genomes aim to address the abundance of genome structural variation within species for improved genomic analyses. New pan-genome, constructed from de novo genome assemblies of 25 diverse melon (Cucumis melo L.) accessions is integrated with a half-diallel derived set of 300 F2 populations representing all possible parental combinations. The potential of this unified multi-parental trait dissection framework for melon genetics and breeding is presented.

SUMMARYLinking genotype with phenotype is a fundamental goal in biology and requires robust data for both. Recent advances in plant‐genome sequencing have expedited comparisons among multiple‐related individuals. The abundance of structural genomic within‐species variation that has been discovered indicates that a single reference genome cannot represent the complete sequence diversity of a species, leading to the expansion of the pan‐genome concept. For high‐resolution forward genetics, this unprecedented access to genomic variation should be paralleled and integrated with phenotypic characterization of genetic diversity. We developed a multi‐parental framework for trait dissection in melon (Cucumis melo), leveraging a novel pan‐genome constructed for this highly variable cucurbit crop. A core subset of 25 diverse founders (MelonCore25), consisting of 24 accessions from the two widely cultivated subspecies of C. melo, encompassing 12 horticultural groups, and 1 feral accession was sequenced using a combination of short‐ and long‐read technologies, and their genomes were assembled de novo. The construction of this melon pan‐genome exposed substantial variation in genome size and structure, including detection of ~300 000 structural variants and ~9 million SNPs. A half‐diallel derived set of 300 F2 populations, representing all possible MelonCore25 parental combinations, was constructed as a framework for trait dissection through integration with the pan‐genome. We demonstrate the potential of this unified framework for genetic analysis of various melon traits, including rind color intensity and pattern, fruit sugar content, and resistance to fungal diseases. We anticipate that utilization of this integrated resource will enhance genetic dissection of important traits and accelerate melon breeding.

SUMMARY Linking genotype with phenotype is a fundamental goal in biology and requires robust data for both. Recent advances in plant‐genome sequencing have expedited comparisons among multiple‐related individuals. The abundance of structural genomic within‐species variation that has been discovered indicates that a single reference genome cannot represent the complete sequence diversity of a species, leading to the expansion of the pan‐genome concept. For high‐resolution forward genetics, this unprecedented access to genomic variation should be paralleled and integrated with phenotypic characterization of genetic diversity. We developed a multi‐parental framework for trait dissection in melon (Cucumis melo), leveraging a novel pan‐genome constructed for this highly variable cucurbit crop. A core subset of 25 diverse founders (MelonCore25), consisting of 24 accessions from the two widely cultivated subspecies of C. melo, encompassing 12 horticultural groups, and 1 feral accession was sequenced using a combination of short‐ and long‐read technologies, and their genomes were assembled de novo. The construction of this melon pan‐genome exposed substantial variation in genome size and structure, including detection of ~300 000 structural variants and ~9 million SNPs. A half‐diallel derived set of 300 F2 populations, representing all possible MelonCore25 parental combinations, was constructed as a framework for trait dissection through integration with the pan‐genome. We demonstrate the potential of this unified framework for genetic analysis of various melon traits, including rind color intensity and pattern, fruit sugar content, and resistance to fungal diseases. We anticipate that utilization of this integrated resource will enhance genetic dissection of important traits and accelerate melon breeding.

Heterosis, the superiority of hybrids over their parents, is a major genetic force associated with plant fitness and crop yield enhancement. We investigated root-mediated yield heterosis in melons (Cucumis melo) by characterizing a common variety grafted onto 190 hybrid rootstocks, resulting from crossing 20 diverse inbreds in a diallel-mating scheme. Hybrid rootstocks improved yield by more than 40% compared with their parents, and the best hybrid yield outperformed the reference commercial variety by 65% under both optimal and minimal irrigation treatments. To characterize the genetics of underground heterosis we conducted whole genome re-sequencing of the 20 founder lines, and showed that parental genetic distance was no predictor for the level of heterosis. Through inference of the 190 hybrid genotypes from their parental genomes, followed by genome-wide association analysis, we mapped multiple quantitative trait loci for root-mediated yield. Yield enhancement of the four best-performing hybrid rootstocks was validated in multiple experiments with four different scion varieties. Our grafting approach is complementary to the common roots genetic approach that focuses mainly on variation in root system architecture, and is a step towards discovery of candidate genes involved in root function and yield enhancement.

Although light is the driving force of photosynthesis, excessive light can be harmful. One of the main processes that limits photosynthesis is photoinhibition, the process of light-induced photodamage. When the absorbed light exceeds the amount that is dissipated by photosynthetic electron flow and other processes, damaging radicals are formed that mostly inactivate photosystem II (PSII). Damaged PSII must be replaced by a newly repaired complex in order to preserve full photosynthetic activity. Chlorella ohadii is a green microalga, isolated from biological desert soil crusts, that thrives under extreme high light and is highly resistant to photoinhibition. Therefore, C. ohadii is an ideal model for studying the molecular mechanisms underlying protection against photoinhibition. Comparison of the thylakoids of C. ohadii cells that were grown under low light versus extreme high light intensities found that the alga employs all three known photoinhibition protection mechanisms: (i) massive reduction of the PSII antenna size; (ii) accumulation of protective carotenoids; and (iii) very rapid repair of photodamaged reaction center proteins. This work elucidated the molecular mechanisms of photoinhibition resistance in one of the most light-tolerant photosynthetic organisms, and shows how photoinhibition protection mechanisms evolved to marginal conditions, enabling photosynthesis-dependent life in severe habitats.

Ornithogalum dubium is a popular ornamental monocot native to South Africa with flower colors ranging from pure white to deep orange. Gene editing based on the CRISPR/Cas9 system has recently been shown to hold potential for color improvement in ornamental flower crops. To apply this approach to Ornithogalum color manipulation, genomic or transcriptomic data must first be collected. Here, cDNA libraries of O. dubium leaves and flowers were constructed and sequenced using the Illumina HiSeq 2500. Over 155 million 100-bp paired-end reads were assembled into a transcriptome database of 360,689 contigs, of which 18,660 contigs were differentially expressed between leaves and flowers. Carotenoids are the main pigment imparting spectrum of orange hues to O. dubium flowers. By querying our database, we identified a total of 16 unique transcripts (unigenes) predicted to be involved in the carotenoid biosynthesis pathway of Ornithogalum. Combining carotenoid profiles, we further inferred several key unigenes responsible for floral coloration and accumulation in O. dubium, of which the gene LCYB/comp146645_c0 was found as a suitable target to generate potentially red flower varieties of O. dubium. Our research thus provides a framework for the application of CRISPR/Cas9 technology to improve this ornamental crop.

SUMMARYPGR5‐LIKE PHOTOSYNTHETIC PHENOTYPE1 (PGRL1) regulates photosystem I cyclic electron flow which transiently activates non‐photochemical quenching at the onset of light. Here, we show that a disulfide‐based mechanism of PGRL1 regulated this process in vivo at the onset of low light levels. We found that PGRL1 regulation depended on active formation of key regulatory disulfides in the dark, and that PGR5 was required for this activity. The disulfide state of PGRL1 was modulated in plants by counteracting reductive and oxidative components and reached a balanced state that depended on the light level. We propose that the redox regulation of PGRL1 fine‐tunes a timely activation of photosynthesis at the onset of low light.

In plants the oxidative cleavage of carotenoid substrates produces volatile apocarotenoids, including β-ionone, 6-methyl-5-hepten-2-ol, and α-ionone; these compounds are important in herbivore-plant communication. Combined chemical, biochemical, and molecular studies were conducted to evaluate the differential accumulation of carotenoids and volatile apocarotenoids during the development of pollinated and parthenocarpic fig fruits. Pollinated fig fruits showed less emission of apocarotenoid volatiles than the parthenocarpic figs, while in the case of carotenoid pigments, pollinated figs manifested higher accumulation. The apocarotenoids, 6-methyl-5-hepten-2-ol and β-cyclogeraniol, showed a marked increase after the two weeks of hand-pollination in pollinated and parthenocarpic figs; but afterwards these volatile levels decreased during further fruit development. In addition, we report a transcriptome-based identification and functional characterization of the carotenoid cleavage dioxygenase (FcCCD) genes. These genes were overexpressed in Escherichia coli strains previously engineered to produce different carotenoids. The recombinant FcCCD1A enzyme showed specificity for the 9,10 (9',10') double bond position of cyclic carotenoids to generate α-ionone and β-ionone, while FcCCD1B cleaved lycopene and an acyclic moiety of δ-carotene, producing 6-methyl-5-hepten-2-one. The qRT-PCR analysis of FcCCD genes revealed differential gene expression during fig fruit development. Our results suggest a role for the FcCCD1genes in apocarotenoid biosynthesis in fig fruits.

To identify factors governing peel-color development in mandarins, we examined carotenoid content and composition and the expression of carotenoid-related genes during four stages of ripening (i.e., green, breaker, yellow, and orange) in two varieties: 'Ora', which has orange fruit, and 'Shani', which has orange-reddish fruit. The two varieties had different carotenoid compositions, and 'Shani' had a significantly higher level of total carotenoid pigments. 'Shani' was rich in the deep orange β-cryptoxanthin and the orange-reddish β-citraurin, whereas 'Ora' was rich in the orange violaxanthin. RNA-Seq analysis revealed significantly greater expression of the carotenoid-biosynthesis genes PSY, βLCY, βCHX, and CCD4b, as well as MEP-pathway genes and several ethylene-biosynthesis and -signaling genes in 'Shani' fruit. In contrast, the expression levels of genes involved in the synthesis of α-branch carotenoids (i.e., εLCY and εCHX) and ZEP, which is involved in the formation of violaxanthin, were significantly higher in the 'Ora' fruit.

AbstractThe sections in this article areIntroductionThe Multifunctional Cell WallTechnical and Conceptual Challenges in Defining the Cell Wall Proteome, or SecretomeApproaches for Plant Cell Wall Protein Extraction and IdentificationComputational Prediction of the Plant SecretomeFunctional Screens for Extracellular ProteinsContamination and Validation of the Cell Wall ProteomeConclusions

Dormancy release is greatly affected by chilling unit (CU) accumulation. Lack of CU has a major impact on spring vegetative budbreak (VB). To understand the genetic mechanism governing the chilling requirement (CR), we conducted a QTL analysis of VB date in F-1 population, derived from a cross between Spadona' (low CR) and Harrow Sweet' (high CR). Using a unique methodology of tree mobility, replicates of the same genotypes were exposed during the winter, over two consecutive years, to climates that differ greatly in their CU and to the same heat conditions to induce VB, in order to evaluate CR genetic impact and to distinguish it from the heat factor. Broad-sense heritability within locations ranged from 0.62 to 0.66. Due to a strong impact of GxE interaction, it was reduced to 0.46 for the overall mean. We examined the previously discovered apple QTLs detected in linkage groups (LG) 9 and 8, based on the synteny between the species. Our analysis confirms significant QTLs in LG8 (R-2=12%-24%) and LG9 (R-2=20%-38%) for all locations and years

Introduction - Carotenoids are tetraterpene pigments which have a major impact on fruit color and nutritional value. Very little is known about the carotenoids' profile and the extent of its variation within different apricot (Prunus armeniaca) accessions. Materials and methods - We analyzed carotenoid content and composition, as well as color criteria, of fruit from 113 different apricot accessions from the Newe Ya'ar germplasm collection in Israel. Results and discussion - Apricot fruit contains a unique profile of carotenoids consisting mainly of beta-carotene, phytoene and phytofluene, and small amounts of other intermediates of the biosynthesis pathway, including cis-lycopene. The different accessions show great variability in total carotenoid content (5-95 mu g g(-1) fresh weight) as well as in carotenoid composition. The percentage of beta-carotene, phytoene and phytofluene varies between 2 to 67%, 6 to 59% and 12 to 44% respectively. Conclusion - Our findings highlight apricot as one of the richest natural sources of the colorless carotenoids phytoene and phytofluene, whose health benefits were recently noted. The distinctive carotenoid profile alongside the high diversity in fruit carotenoid composition and content among apricot varieties can assist future breeding programs and may help in understanding the factors contributing to color and nutritional traits of apricot fruit.

Most commercial apple cultivars have high to medium chilling requirements and consequently are not grown in regions with warm winters. Furthermore, global climate changes raise the concern that some regions where apples are currently being produced will become unsuitable in the future. Therefore, mapping and understanding the factors governing chilling requirements are important goals towards the breeding of new apple varieties. In this study, we characterized 73 apple accessions: old local accessions, modern cultivars, and selected hybrids, all grown in the Newe Ya’ar germplasm collection in Israel under moderate winter conditions. We measured the time of vegetative bud-break as an indicator of chilling requirements and genotyped the accessions for known genetic markers and for markers we developed by re-sequencing the genome of ‘Anna’, a cultivar with very low chilling requirements. Our results show that while most of the accessions that were characterized as having early bud-break are genetically different from each other, they all share a unique haplotype in a region of ∼190 kb, within a previously identified QTL for bud-break time, on chromosome 9. The alleles in the early bud-break-associated haplotype were not found in any of the late accessions tested, suggesting that the causative difference leading to the variation in bud-break time lays within or near this region, and that there is a common ancestor carrying early bud-break trait of the early accessions tested. Moreover, the markers of the unique haplotype can serve as genetic markers to accelerate the breeding of apple cultivars better adapted to warm climates.

Plant cuticles on outer fruit and leaf surfaces are natural macromolecular composites of waxes and polyesters that ensure mechanical integrity and mitigate environmental challenges. They also provide renewable raw materials for cosmetics, packaging, and coatings. To delineate the structural framework and flexibility underlying the versatile functions of cutin biopolymers associated with polysaccharide-rich cell-wall matrices, solid-state NMR spectra and spin relaxation times were measured in a tomato fruit model system, including different developmental stages and surface phenotypes. The hydrophilic-hydrophobic balance of the cutin ensures compatibility with the underlying polysaccharide cell walls; the hydroxy fatty acid structures of outer epidermal cutin also support deposition of hydrophobic waxes and aromatic moieties while promoting the formation of cell-wall cross-links that rigidify and strengthen the cuticle composite during fruit development. Fruit cutin-deficient tomato mutants with compromised microbial resistance exhibit less efficient local and collective biopolymer motions, stiffening their cuticular surfaces and increasing their susceptibility to fracture.

Bay laurel (Laurus nobilis L.) is an agriculturally important tree used in food, drugs, and the cosmetics industry. Many of the health beneficial properties of bay laurel are due to volatile terpene metabolites that they contain, including various norisoprenoids. Despite their importance, little is known about the norisoprenoid biosynthesis in Laurus nobilis fruits. We found that the volatile norisoprenoids 6-methyl-5-hepten-2-one, pseudoionone, and β-ionone accumulated in Laurus nobilis fruits in a pattern reflecting their carotenoid content. A full-length cDNA encoding a potential carotenoid cleavage dioxygenase (LnCCD1) was isolated. The LnCCD1 gene was overexpressed in Escherichia coli, and recombinant protein was assayed for its cleavage activity with an array of carotenoid substrates. The LnCCD1 protein was able to cleave a variety of carotenoids at the 9,10 (9',10') and 5,6 (5',6') positions to produce 6-methyl-5-hepten-2-one, pseudoionone, β-ionone, and α-ionone. Our results suggest a role for LnCCD1 in Laurus nobilis fruit flavor biosynthesis.