Summary
Combined quantitative trait loci (QTL) and expression‐QTL (eQTL) mapping analysis was performed to identify genetic factors affecting melon (Cucumis melo) fruit quality, by linking genotypic, ...metabolic and transcriptomic data from a melon recombinant inbred line (RIL) population. RNA sequencing (RNA‐Seq) of fruit from 96 RILs yielded a highly saturated collection of > 58 000 single‐nucleotide polymorphisms, identifying 6636 recombination events that separated the genome into 3663 genomic bins. Bin‐based QTL analysis of 79 RILs and 129 fruit‐quality traits affecting taste, aroma and color resulted in the mapping of 241 QTL. Thiol acyltransferase (CmThAT1) gene was identified within the QTL interval of its product, S‐methyl‐thioacetate, a key component of melon fruit aroma. Metabolic activity of CmThAT1‐encoded protein was validated in bacteria and in vitro. QTL analysis of flesh color intensity identified a candidate white‐flesh gene (CmPPR1), one of two major loci determining fruit flesh color in melon. CmPPR1 encodes a member of the pentatricopeptide protein family, involved in processing of RNA in plastids, where carotenoid and chlorophyll pigments accumulate. Network analysis of > 12 000 eQTL mapped for > 8000 differentially expressed fruit genes supported the role of CmPPR1 in determining the expression level of plastid targeted genes. We highlight the potential of RNA‐Seq‐based QTL analysis of small to moderate size, advanced RIL populations for precise marker‐assisted breeding and gene discovery. We provide the following resources: a RIL population genotyped with a unique set of SNP markers, confined genomic segments that harbor QTL governing 129 traits and a saturated set of melon eQTLs.
Significance Statement
Fruit‐quality traits of melon were studied by linking genotypic, metabolic and transcriptomic data from a recombinant inbred line population. High resolution RNA‐Seq‐based quantitative trait loci and expression‐QTL analyses enabled the identification of two genes that affect aroma and color.
Color is an important determinant of pomegranate fruit quality and commercial value. To understand the genetic factors controlling color in pomegranate, chemical, molecular and genetic ...characterization of a "white" pomegranate was performed. This unique accession is lacking the typical pomegranate color rendered by anthocyanins in all tissues of the plant, including flowers, fruit (skin and arils) and leaves. Steady-state gene-expression analysis indicated that none of the analyzed "white" pomegranate tissues are able to synthesize mRNA corresponding to the PgLDOX gene (leucoanthocyanidin dioxygenase, also called ANS, anthocyanidin synthase), which is one of the central structural genes in the anthocyanin-biosynthesis pathway. HPLC analysis revealed that none of the "white" pomegranate tissues accumulate anthocyanins, whereas other flavonoids, corresponding to biochemical reactions upstream of LDOX, were present. Molecular analysis of the "white" pomegranate revealed the presence of an insertion and an SNP within the coding region of PgLDOX. It was found that the SNP does not change amino acid sequence and is not fully linked with the "white" phenotype in all pomegranate accessions from the collection. On the other hand, genotyping of pomegranate accessions from the collection and segregating populations for the "white" phenotype demonstrated its complete linkage with the insertion, inherited as a recessive single-gene trait. Taken together, the results indicate that the insertion in PgLDOX is responsible for the "white" anthocyanin-less phenotype. These data provide the first direct molecular, genetic and chemical evidence for the effect of a natural modification in the LDOX gene on color accumulation in a fruit-bearing woody perennial deciduous tree. This modification can be further utilized to elucidate the physiological role of anthocyanins in protecting the tree organs from harmful environmental conditions, such as temperature and UV radiation.
Demand for almond is in rise, due to its desired flavor and beneficial health-related properties. Roasting is the most common practice in almond processing, improving aroma and texture, as well as ...nutritional properties. “Shefa,” a new Israeli almond variety, was recently introduced; however, no information is available regarding its phytochemical composition and response to roasting. The aim of the current work was to characterize its health-related composition, in addition to describe its response to moderate roasting conditions, including temperature and duration. For this, “Shefa” almonds were roasted at 100°C, 125°C, 150°C, and 175°C for 15 and 25 min. and compared to raw unroasted kernels. Health and nutritional quality parameters included antioxidant capacity (DPPH, FRAP, and ABTS; spectrophotometric), total polyphenol, tocopherol and phytosterol content (spectrophotometric), and fatty acid profiling (GC-MS). Our results demonstrate that “Shefa” variety phytochemical composition resembles that of other Mediterranean almond varieties, e.g. Italian and Turkish are high in tocopherols and phytosterols, with average polyphenol content. Its antioxidant activity was 19.11, 25.56, and 22.18 mg TE/100 g DW for DPPH, FRAP, and ABTS, respectively. In addition, under the roasting conditions evaluated in the current work, “Shefa” almonds presented an improved health-related composition, with levels of antioxidant capacity, polyphenols, tocopherols, and phytosterols increasing with temperature and duration. “Shefa” response to roasting resembled that of other almond varieties evaluated under comparable roasting conditions. To conclude, the newly introduced Israeli almond “Shefa” has a healthy and nutritionally beneficial phytochemical composition, which improves with moderate roasting. Further investigation is needed to profile its polyphenol and tocopherol composition, to provide additional valuable insights into its phytochemical composition.
A number of molecular marker linkage maps have been developed for melon (Cucumis melo L.) over the last two decades. However, these maps were constructed using different marker sets, thus, making ...comparative analysis among maps difficult. In order to solve this problem, a consensus genetic map in melon was constructed using primarily highly transferable anchor markers that have broad potential use for mapping, synteny, and comparative quantitative trait loci (QTL) analysis, increasing breeding effectiveness and efficiency via marker-assisted selection (MAS).
Under the framework of the International Cucurbit Genomics Initiative (ICuGI, http://www.icugi.org), an integrated genetic map has been constructed by merging data from eight independent mapping experiments using a genetically diverse array of parental lines. The consensus map spans 1150 cM across the 12 melon linkage groups and is composed of 1592 markers (640 SSRs, 330 SNPs, 252 AFLPs, 239 RFLPs, 89 RAPDs, 15 IMAs, 16 indels and 11 morphological traits) with a mean marker density of 0.72 cM/marker. One hundred and ninety-six of these markers (157 SSRs, 32 SNPs, 6 indels and 1 RAPD) were newly developed, mapped or provided by industry representatives as released markers, including 27 SNPs and 5 indels from genes involved in the organic acid metabolism and transport, and 58 EST-SSRs. Additionally, 85 of 822 SSR markers contributed by Syngenta Seeds were included in the integrated map. In addition, 370 QTL controlling 62 traits from 18 previously reported mapping experiments using genetically diverse parental genotypes were also integrated into the consensus map. Some QTL associated with economically important traits detected in separate studies mapped to similar genomic positions. For example, independently identified QTL controlling fruit shape were mapped on similar genomic positions, suggesting that such QTL are possibly responsible for the phenotypic variability observed for this trait in a broad array of melon germplasm.
Even though relatively unsaturated genetic maps in a diverse set of melon market types have been published, the integrated saturated map presented herein should be considered the initial reference map for melon. Most of the mapped markers contained in the reference map are polymorphic in diverse collection of germplasm, and thus are potentially transferrable to a broad array of genetic experimentation (e.g., integration of physical and genetic maps, colinearity analysis, map-based gene cloning, epistasis dissection, and marker-assisted selection).
Pomegranate is a valuable crop that is grown commercially in many parts of the world. Wild species have been reported from India, Turkmenistan and Socotra. Pomegranate fruit has a variety of ...health-beneficial qualities. However, despite this crop's importance, only moderate effort has been invested in studying its biochemical or physiological properties or in establishing genomic and genetic infrastructures. In this study, we reconstructed a transcriptome from two phenotypically different accessions using 454-GS-FLX Titanium technology. These data were used to explore the functional annotation of 45,187 fully annotated contigs. We further compiled a genetic-variation resource of 7,155 simple-sequence repeats (SSRs) and 6,500 single-nucleotide polymorphisms (SNPs). A subset of 480 SNPs was sampled to investigate the genetic structure of the broad pomegranate germplasm collection at the Agricultural Research Organization (ARO), which includes accessions from different geographical areas worldwide. This subset of SNPs was found to be polymorphic, with 10.7% loci with minor allele frequencies of (MAF<0.05). These SNPs were successfully used to classify the ARO pomegranate collection into two major groups of accessions: one from India, China and Iran, composed of mainly unknown country origin and which was more of an admixture than the other major group, composed of accessions mainly from the Mediterranean basin, Central Asia and California. This study establishes a high-throughput transcriptome and genetic-marker infrastructure. Moreover, it sheds new light on the genetic interrelations between pomegranate species worldwide and more accurately defines their genetic nature.
Almond
(Mill.) D. A. Webb is a major deciduous fruit tree crop worldwide. During dormancy, under warmer temperatures and inadequate chilling hours, the plant metabolic activity increases and may ...lead to carbohydrate deficiency.
(Olivier) Meikle is a bushy wild almond species known for its green, unbarked stem, which stays green even during the dormancy period. Our study revealed that
green stems assimilate significantly high rates of CO
during the winter as compared to
cv. Um el Fahem (U.E.F.) and may improve carbohydrate status throughout dormancy. To uncover the genetic inheritance and mechanism behind the
stem photosynthetic capability (SPC), a segregated F1 population was generated by crossing
to U.E.F. Both parent's whole genome was sequenced, and SNP calling identified 4,887 informative SNPs for genotyping. A robust genetic map for U.E.F. and
was constructed (971 and 571 markers, respectively). QTL mapping and association study for the SPC phenotype revealed major QTL log of odd (LOD) = 20.8 on chromosome 7 and another minor but significant QTL on chromosome 1 (LOD = 3.9). As expected, the
allele in the current loci significantly increased the SPC phenotype. Finally, a list of 64 candidate genes was generated. This work sets the stage for future research to investigate the mechanism regulating the SPC trait, how it affects the tree's physiology, and its importance for breeding new cultivars better adapted to high winter temperatures.
Anthocyanins are important dietary and health-promoting substances present in high quantities in the peel and arils of the pomegranate (
L.) fruit. Yet, there is a high variation in the content of ...anthocyanin among different pomegranate varieties. The 'Black' pomegranate variety (P.G.127-28) found in Israel contains exceptionally high levels of anthocyanins in its fruit peel which can reach up to two orders of magnitude higher content as compared to that of other pomegranate varieties' peel anthocyanins. Biochemical analysis reveals that delphinidin is highly abundant in the peel of 'Black' variety. The pattern of anthocyanin accumulation in the fruit peel during fruit development of 'Black' variety differs from that of other pomegranates. High anthocyanin levels are maintained during all developmental stages. Moreover, the accumulation of anthocyanin in the fruit peel of 'Black' variety is not dependent on light. Genetic analysis of an F
population segregating for the "black" phenotype reveals that it is determined by a single recessive gene. Genetic mapping of the F
population using single nucleotide polymorphism (SNP) markers identified few markers tightly linked to the "black" phenotype. Recombination analysis of the F
population and F
populations narrowed the "black" trait to an area of 178.5 kb on the draft genome sequence of pomegranate
. 'Dabenzi.' A putative
(
) gene is located in this area. Only pomegranate varieties displaying the "black" trait carry a base pair deletion toward the end of the gene, causing a frame shift resulting in a shorter protein. We propose that this mutation in the
gene is responsible for the different anthocyanin composition and high anthocyanin levels of the "black" trait in pomegranate.
The pomegranate (
L.) is a deciduous fruit tree that grows worldwide. However, there are variants, which stay green in mild winter conditions and are determined evergreen. The evergreen trait is of ...commercial and scientific importance as it extends the period of fruit production and provides opportunity to identify genetic functions that are involved in sensing environmental cues. Several different evergreen pomegranate accessions from different genetic sources grow in the Israeli pomegranate collection. The leaves of deciduous pomegranates begin to lose chlorophyll during mid of September, while evergreen accessions continue to generate new buds. When winter temperature decreases 10°C, evergreen variants cease growing, but as soon as temperatures arise budding starts, weeks before the response of the deciduous varieties. In order to understand the genetic components that control the evergreen/deciduous phenotype, several segregating populations were constructed, and high-resolution genetic maps were assembled. Analysis of three segregating populations showed that the evergreen/deciduous trait in pomegranate is controlled by one major gene that mapped to linkage group 3. Fine mapping with advanced F3 and F4 populations and data from the pomegranate genome sequences revealed that a gene encoding for a putative and unique MADS transcription factor (
) is responsible for the evergreen trait. Ectopic expression of
in Arabidopsis generated small plants and early flowering. The deduced protein of
includes eight glutamines (polyQ) at the N-terminus. Three-dimensional protein model suggests that the polyQ domain structure might be involved in DNA binding of PgMADS. Interestingly, all the evergreen pomegranate varieties contain a mutation within the polyQ that cause a stop codon at the N terminal. The polyQ domain of PgPolyQ-MADS resembles that of the ELF3 prion-like domain recently reported to act as a thermo-sensor in Arabidopsis, suggesting that similar function could be attributed to PgPolyQ-MADS protein in control of dormancy. The study of the evergreen trait broadens our understanding of the molecular mechanism related to response to environmental cues. This enables the development of new cultivars that are better adapted to a wide range of climatic conditions.
The sweet melon fruit is characterized by a metabolic transition during its development that leads to extensive accumulation of the disaccharide sucrose in the mature fruit. While the biochemistry of ...the sugar metabolism pathway of the cucurbits has been well studied, a comprehensive analysis of the pathway at the transcriptional level allows for a global genomic view of sugar metabolism during fruit sink development. We identified 42 genes encoding the enzymatic reactions of the sugar metabolism pathway in melon. The expression pattern of the 42 genes during fruit development of the sweet melon cv Dulce was determined from a deep sequencing analysis performed by 454 pyrosequencing technology, comprising over 350,000 transcripts from four stages of developing melon fruit flesh, allowing for digital expression of the complete metabolic pathway. The results shed light on the transcriptional control of sugar metabolism in the developing sweet melon fruit, particularly the metabolic transition to sucrose accumulation, and point to a concerted metabolic transition that occurs during fruit development.
Zucchini yellow mosaic virus (ZYMV; potyviridae) represents a major pathogen of Cucurbitaceae crops. ZYMV resistance in melon PI 414723 is conditioned by a dominant allele at the Zym locus. This ...resistant accession restricts viral spread and does not develop mosaic symptoms, but necrosis sometimes develops in response to inoculation. In previous studies, Zym has been mapped to linkage group II of the melon genetic map. In the present study, positional cloning of the locus was undertaken, starting from the CM-AG36 SSR marker at approximately 2 cm distance. We utilized five mapping populations that share the same resistant parent, PI 414723, and analyzed a total of 1630 offspring, to construct a high-resolution genetic map of the Zym locus. Two melon BAC libraries were used for chromosome walking and for developing new markers closer to the resistance gene by BAC-end sequencing. A BAC contig was constructed, and we identified a single BAC clone, from the ZYMV susceptible genotype MR-1, that physically encompasses the resistance gene. A second clone was isolated from another susceptible genotype, WMR 29, and the two clones were fully sequenced and annotated. Additional markers derived from the sequenced region delimited the region to 17.6 kb of a sequence that harbors a NAC-like transcription factor and, depending on the genotype, either two or three R-gene homologs with a CC-NBS-LRR structure. Mapping was confirmed by saturating the map with SNP markers using a single mapping population. The same region was amplified and sequenced also in the ZYMV resistant genotype PI 414723. Because numerous polymorphic sites were noted between genotypes, we could not associate resistance with a specific DNA polymorphism; however, this study enables molecular identification of Zym and paves the way to functional studies of this important locus.
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