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Genetic analysis revealed that CmCLV3 is a candidate gene for the variation in melon carpel number. Carpel number (CN) is an important trait in melon. Three-CN melon fruit is oval, while 5-CN melon fruit has a round or flat shape. Herein, a genetic analysis of a population in which the CN locus was segregated indicated that 3-CN is controlled by a major dominant effective gene. Bulked segregant analysis and initial linkage mapping placed the CN locus in a 6.67 Mb region on chromosome 12, and it was narrowed to 882.19 kb with molecular markers and recombinant plants. Fine mapping with a large F

Vitamin A deficiency is a worldwide public nutrition problem, and β-carotene is the precursor for vitamin A synthesis. Watermelon with golden flesh (gf, due to accumulated abundance of β-carotene) is an important germplasm resource. In this study, a genetic analysis of gf segregating populations indicated that gf was controlled by a single recessive gene. BSA-seq and an initial linkage analysis placed the gf locus in a 290-Kb region on watermelon chromosome 1. Further fine mapping in a large population with over 1,000 F2 plants narrowed this region to 39.08 Kb harboring two genes, Cla97C01G008760 and Cla97C01G008770, which encode phytoene synthase (ClPsy1) and GATA zinc finger domain-containing protein, respectively. Gene sequence alignment and expression analysis between parental lines revealed Cla97C01G008760 as the best possible candidate gene for gf trait. Nonsynonymous SNP mutations in the first exon of ClPsy1 between parental lines cosegregated with the gf trait only among individuals in the genetic population but were not related to flesh color in natural watermelon panels. Promoter sequence analysis of 26 watermelon accessions revealed two SNPs in the cis-acting element sequences corresponding to MYB and MYC2 transcription factors. RNA-seq data and qRT-PCR verification showed that two MYBs and one MYC2 exhibited expression trends similar as ClPsy1 in the parental lines, which may thus play roles in the regulation of ClPsy1 expression. Our research findings indicate that the gf trait is determined not only by ClPsy1 but also by ClLCYB, ClCRTISO and ClNCED7, which play important roles in β-carotene accumulation in watermelon flesh.

The color of watermelon flesh is an important trait determined by a series of carotenoids. Herein, we used Cream of Saskatchewan (pale yellow flesh) and PI 186490 (white flesh) as parental materials for an F2 segregation and initial mapping using the bulked segregant analysis sequencing (BSA-seq) strategy. The BSA results revealed a flesh color-related QTL that spans approximately 2.45 Mb on chromosome 6. This region was preliminarily positioned in a 382-kb segment, and then narrowed down into a 66.8-kb segment with 1260 F2 individuals. A total of nine candidate genes were in the fine mapping interval, but only Cla007528 (encoding chlorophyllase) had non-synonymous mutations and was significantly expressed between the parental materials throughout flesh development. We also checked the expression patterns of the carotenoid metabolic pathway genes based on RNA-seq data and qRT-PCR validation. Three genes in the xanthophyll cycle (CICHYB, CINCED-1 and CINCED-7) exhibited differential expression patterns between the two parental lines at different flesh color formation stages. ClPSY1, CIPDS, CIZDS, CICHXE, CICRTISO and CILCYB also exhibited clearly different expression patterns accompanied by carotenoid accumulation.

Stigma color plays an important role in pollination. In nature, melon (Cucumis melo L.) stigmas are either yellow or green; however, a review of the literature found no report on how stigma color affects pollination and fruit development in melon. Here, we used an F2 melon population derived from a cross between ‘MR-1’ (P1, with green stigmas) and ‘M1–32’ (P2, with yellow stigmas), and performed genetic analysis and mapping. The results of bulked segregant analysis allowed the identification of genetic loci controlling stigma color on chromosomes 6 and 8. An F2 population consisting of 150 individuals was used for initial mapping. A genetic map of 304.17 cM was constructed using 37 cleaved amplified polymorphism sequence (CAPS) markers. We identified one major quantitative trait locus (QTL) and one minor QTL for stigma color. The major QTL GS8.1 was further mapped to a 4.13 cM interval between CAPS markers 8C-10 and 8C-16, which explained 27.04% of the phenotypic variation. In addition, GS6.1 was mapped between E-49 and 6A-7, explaining 18.6% of the phenotypic variation. This study provides a theoretical basis for the fine mapping and cloning of melon genes controlling stigma color.

The plant compact and dwarf growth habit is an important agronomic trait when breeding watermelon (Citrullus lanatus) cultivars because of their reduced vine length, high-density planting, and better land utilization; however, the genetic basis of the dwarf growth habit is not well-known. In this study, the plant population of six generations, P1, P2, F1, F2, BC1P1, and BC1P2, were studied. A genetic segregation analysis demonstrated that dwarfism is mainly controlled by a single recessive Cldw gene. Furthermore, whole-genome sequencing of two distinct watermelon cultivars, W1-1 (P1) and 812 (P2), was performed and preliminarily mapped through a bulked segregant analysis of F2 individuals that revealed the Cldw gene locus on chromosome 9. Two candidate genes, Cla015407 and Cla015408, were discovered at the delimited region of 43.2 kb by fine mapping, and gene annotation exposed that the Cla015407 gene encodes gibberellic acid 3β-hydroxylase protein. In addition, a comparative analysis of gene sequence and cultivars sequences across the reference genome of watermelon revealed the splice site mutation in the intron region of the Cldw gene in dwarf-type cultivar 812. The quantitative real-time polymerase chain reaction exhibited a significantly higher expression of the Cla015407 gene in cultivar W1-1 compared with 812. There was no significant difference in the vine length of both cultivars after gibberellic acid treatment. In brief, our fine mapping demonstrated that Cla015407 is a candidate gene controlling dwarfism of watermelon plants.

Thaumatin-like proteins (TLPs), which belong to pathogenesis-related (PR) protein family 5 (PR5), are involved in plant host defense and various developmental processes. The functions of the TLP family have been extensively discussed in multiple organisms, whereas the detailed information of this family in melon has not been reported yet. In this study, we identified 28 TLP genes in the melon genome and a N-terminal signal peptide was found highly conserved within each member of this family. Phylogeny analysis indicated that TLPs from melon and other plant species were clustered into ten groups. Twelve segmental and seven tandem duplication gene pairs that underwent purifying selection were identified. TLP genes expressed differentially in different tissues/organs, and were significantly induced after Podosphaera xanthii infection. TLPs in breeding line MR-1 tend to express early after pathogen infection compared with cultivar Top Mark. Our study provides a comprehensive understanding of the melon TLP family and demonstrates their potential roles in disease resistance, therefore provides more reference for further research.

The Cucurbitaceae family hosts many economically important fruit vegetables (cucurbits) such as cucumber, melon, watermelon, pumpkin/squash, and various gourds. The cucurbits are probably best known for the diverse fruit sizes and shapes, but little is known about their genetic basis and molecular regulation. Here, we reviewed the literature on fruit size (FS), shape (FSI), and fruit weight (FW) QTL identified in cucumber, melon, and watermelon, from which 150 consensus QTL for these traits were inferred. Genome-wide survey of the three cucurbit genomes identified 253 homologs of eight classes of fruit or grain size/weight-related genes cloned in Arabidopsis, tomato, and rice that encode proteins containing the characteristic CNR (cell number regulator), CSR (cell size regulator), CYP78A (cytochrome P450), SUN, OVATE, TRM (TONNEAU1 Recruiting Motif), YABBY, and WOX domains. Alignment of the consensus QTL with candidate gene homologs revealed widespread structure and function conservation of fruit size/shape gene homologs in cucurbits, which was exemplified with the fruit size/shape candidate genes CsSUN25-26-27a and CsTRM5 in cucumber, CmOFP1a in melon, and ClSUN25-26-27a in watermelon. In cucurbits, the andromonoecy (for 1-aminocyclopropane-1-carboxylate synthase) and the carpel number (for CLAVATA3) loci are known to have pleiotropic effects on fruit shape, which may complicate identification of fruit size/shape candidate genes in these regions. The present work illustrates the power of comparative analysis in understanding the genetic architecture of fruit size/shape variation, which may facilitate QTL mapping and cloning for fruit size-related traits in cucurbits. The limitations and perspectives of this approach are also discussed.

Fruit morphology traits are important commercial traits that directly affect the market value. However, studying the genetic basis of these traits in un-explored botanical groups is a fundamental objective for crop genetic improvement through marker-assisted breeding.In this study, a quantitative trait loci (QTLs) mapping strategy was used for dissecting the genomic regions of fruit linked morphological traits by single nucleotide polymorphism (SNP) based cleaved amplified polymorphism sequence (CAPS) molecular markers. Next-generation sequencing was done for the genomic sequencing of two contrasted melon lines (climacteric and non-climacteric), which revealed 97% and 96% of average coverage over the reference melon genome database, respectively. A total of 57.51% non-synonymous SNPs and 42.49% synonymous SNPs were found, which produced 149 sets of codominant markers with a 24% polymorphism rate. Total 138-FIn short, our illustrated genetic loci are expected to provide the reference insights for fine QTL mapping and candidate gene(s) mining through molecular genetic breeding approaches aimed at developing the new varieties.

The three-seeded pod number is an important trait that positively influences soybean yield. Soybean variety with increased three-seeded pod number contributes to the seed number/plant and higher yield. The candidate genes of the three-seeded pod may be the key for improving soybean yield. In this study, identification and validation of candidate genes for three-seeded pod has been carried out. First, a total of 36 quantitative trait locus (QTL) were detected from the investigation of recombinant inbred lines including 147 individuals derived from a cross between Charleston and Dongning 594 cultivars. Five consensus QTLs were integrated. Second, an introgressed line CSSL-182 carrying the target segment for the trait from the donor parent was selected to verify the consensus QTL based on its phenotype. Third, a secondary group was constructed by backcrossing with CSSL-182, and two QTLs were confirmed. There were a total of 162 genes in the two QTLs. The mining of candidate genes resulted in the annotation of eight genes with functions related to pod and seed sets. Finally, haplotype analysis and quantitative reverse transcriptase real-time PCR were carried to verify the candidate genes. Four of these genes had different haplotypes in the resource group, and the differences in the phenotype were highly significant. Moreover, the differences in the expression of the four genes during pod and seed development were also significant. These four genes were probably related to the development process underlying the three-seeded pod in soybean. Herein, we discuss the past and present studies related to the three-seeded pod trait in soybean.

Homeodomain-leucine zipper (HD-ZIP) transcription factors are one of the plant-specific gene families involved in plant growth and response to adverse environmental conditions. However, little information is available on the HD-ZIP gene family in watermelon. In this study, forty ClHDZs were systemically identified in the watermelon genome, which were subsequently divided into four distinctive subfamilies (I–IV) based on the phylogenetic topology. HD-ZIP members in the same subfamily generally shared similar gene structures and conserved motifs. Syntenic analyses revealed that segmental duplications mainly contributed to the expansion of the watermelon HD-ZIP family, especially in subfamilies I and IV. HD-ZIP III was considered the most conserved subfamily during the evolutionary history. Moreover, expression profiling together with stress-related cis-elements in the promoter region unfolded the divergent transcriptional accumulation patterns under abiotic stresses. The majority (13/23) of ClHDZs in subfamilies I and II were downregulated under the drought condition, e.g., ClHDZ4, ClHDZ13, ClHDZ18, ClHDZ19, ClHDZ20, and ClHDZ35. On the contrary, most HD-ZIP genes were induced by cold and salt stimuli with few exceptions, such as ClHDZ3 and ClHDZ23 under cold stress and ClHDZ14 and ClHDZ15 under the salt condition. Notably, the gene ClHDZ14 was predominantly downregulated by three stresses whereas ClHDZ1 was upregulated, suggesting their possible core roles in response to these abiotic stimuli. Collectively, our findings provide promising candidates for the further genetic improvement of abiotic stress tolerance in watermelon.

The genic male sterility (MS) plays a major role in melon hybrids production, it could reduce the cost of pollination and increase the yield and quality. However, the molecular mechanism underlying genetic male sterility is yet poorly understood. The morphological differences of flower buds of melon were observed showed that the flower buds were tetrad when they were 1 mm stage and monocyte microspore when they were 2 mm stage. Electron microscopy showed that there was significant difference between MS lines and MF (male fertility) lines. In order to detect the global expression of the genes during the melon anther development and association with MS, 12 DEGs (differentially expressed genes) libraries were constructed from the anther of MS and MF in the bud stage with 1 and 2 mm diameter, respectively. A total of 765 DEGs expressed in anther during different developmental stage (MS 1 mm vs. MS 2 mm), 148 and 309 DEGs were found to be related to MS as compared to MF (MS 1 mm vs. MF 1 mm, and MS 2 mm vs. MF 2 mm) at a false discovery rate FDR

Phytoene synthase (PSY) plays an essential role in carotenoid biosynthesis. In this study, three ClPSY genes were identified through the watermelon genome, and their full-length cDNA sequences were cloned. The deduced proteins of the three ClPSY genes were ranged from 355 to 421 amino acid residues. Phylogenetic analysis suggested that the ClPSYs are highly conserved with bottle gourd compared to other cucurbit crops PSY proteins. Variation in ClPSY1 expression in watermelon with different flesh colors was observed; ClPSY1 was most highly expressed in fruit flesh and associated with the flesh color formation. ClPSY1 expression was much lower in the white-fleshed variety than the colored fruits. Gene expression analysis of ClPSY genes in root, stem, leaf, flower, ovary and flesh of watermelon plants showed that the levels of ClPSY2 transcripts found in leaves was higher than other tissues; ClPSY3 was dominantly expressed in roots. Functional complementation assays of the three ClPSY genes suggested that all of them could encode functional enzymes to synthesize the phytoene from Geranylgeranyl Pyrophosphate (GGPP). Some of the homologous genes clustered together in the phylogenetic tree and located in the synteny chromosome region seemed to have similar expression profiles among different cucurbit crops. The findings provide a foundation for watermelon flesh color breeding with regard to carotenoid synthesis and also provide an insight for the further research of watermelon flesh color formation.

Two genes for resistance to Podosphaera xanthii race 1 in melon were identified on chromosomes 10 and 12 of the Cucumis melo cultivar MR-1. Cucumis melo L. is an economically important crop, the production of which is threatened by the prevalence of melon powdery mildew (PM) infections. We herein utilized the MR-1 (P

Fruit pedicel length (FPL) is among the most important traits associated with melon (Cucumis melo L.), impacting their appearance and management. Melon pedicels are also utilized in the preparation of certain traditional Chinese medicine formulations. Herein, we analyzed a melon parental lines with long and short pedicles (M125 and X055, respectively), and found that the primary differences between these lines included changes in both cell number and cytokinin content in the fruit pedicel. This study is the first to report that the CmFpl3.1 is associated with the regulation of melon pedicel length, and we were able to narrow this locus to an 89 Kb region containing 10 genes on chromosome 3 via bulk segregant analysis (BSA) and quantitative trait loci (QTL) analysis. Microscopy, qPCR, functional annotation, and gene sequence alignment studies further led us to identify MELO3C010972, which encodes cytokinin oxidase, as a candidate gene of CmFpl3.1 likely to regulate melon FPL. Furthermore, the concomitant cleave amplified polymorphic sequence (CAPS) marker genotyping data sets from 95 germplasm accessions strongly supported a role for MELO3C010972 as a regulator of FPL variability. In conclusion, our results offer new insights regarding the mechanistic basis for FPL determination, offering a foundation for future breeding programs or efforts to understand the mechanisms governing melon FPL development.

Cucurbits (Cucurbitaceae family) include many economically important fruit vegetable crops such as watermelon, pumpkin/squash, cucumber, and melon. Seed size (SS) is an important trait in cucurbits breeding, which is controlled by quantitative trait loci (QTL). Recent advances have deciphered several signaling pathways underlying seed size variation in model plants such as Arabidopsis and rice, but little is known on the genetic basis of SS variation in cucurbits. Here we conducted literature review on seed size QTL identified in watermelon, pumpkin/squash, cucumber and melon, and inferred 14, 9 and 13 consensus SS QTL based on their physical positions in respective draft genomes. Among them, four from watermelon (ClSS2.2, ClSS6.1, ClSS6.2, and ClSS8.2), two from cucumber (CsSS4.1 and CsSS5.1), and one from melon (CmSS11.1) were major-effect, stable QTL for seed size and weight. Whole genome sequence alignment revealed that these major-effect QTL were located in syntenic regions across different genomes suggesting possible structural and functional conservation of some important genes for seed size control in cucurbit crops. Annotation of genes in the four watermelon consensus SS QTL regions identified genes that are known to play important roles in seed size control including members of the zinc finger protein and the E3 ubiquitin-protein ligase families. The present work highlights the utility of comparative analysis in understanding the genetic basis of seed size variation, which may help future mapping and cloning of seed size QTL in cucurbits.

Fruit traits influence consumer’s choice and the market worth of melons. Multiple genes controlled the majority of fruit quality traits and these genes are influenced via differential factors. Cleaved amplified polymorphism sequence (CAPS) molecular markers have been validated as efficient tool for detection of numerous SNPs (single nucleotide polymorphisms). In the current study, genetic linkage map was assembled and QTL discovery was conducted for melon quality traits in F2:3 population, generated from crossing of MR-1 and M4-7. SNPs were detected and converted into CAPS markers from retrieved data of high-throughput resequencing. Two parental lines (MR-1 and M4-7) of melon were analyzed and resequenced for assembling the novel CAPS markers. Totally, 91.54% and 90.00% of assembled sequences covered the reference genome of these two distinct parental lines, respectively. Analysis of assembled genomic data revealed total 2,761,801 SNPs and 7741 CAPS locus. Overall, 410 novels CAPS molecular markers were assembled along with 4-restriction endonuclease and 160 CAPS pairs displayed polymorphic bands of predictable length after screening by the use of PCR and process of enzyme digestion, with 39% rate of polymorphism. Genetic linkage map construction was done by genotyping of these novel CAPS markers within F2:3 population and this map totally spanned 2848.68 cM length with 12 chromosomes and 160 CAPS markers. Total 24 minor QTLs were discovered for fruit weight, firmness, flesh firmness, Brix, fruit length, width and shape with different genetic loci on various chromosomal locations. These newly assembled CAPS markers will be valuable and potent for future breeding and genetic schemes in melon.

Powdery mildew (PM) is a catastrophic disease of melon caused by Podosphaera xanthii (Px). However, there is limited information available at the molecular level regarding how the melon plant develops defense mechanisms against this pathogen. We studied melon mRNA transcripts from the resistant MR-1 and susceptible Topmark cultivars through comparative transcriptome analysis. Leaves inoculated with spores of P. xanthii were collected from 0 to 168 hpi to identify the candidate regulators for resistance to PM at different infection stages. We identified 1888 and 2842 melon genes in MR-1 and Topmark, respectively, that were differentially expressed during the plant-pathogen interaction. Gene function analysis of these differentially expressed genes (DEGs) classified them into seven functional groups: pathogen recognition, signal transduction, transcription factors (TFs), phytoalexin biosynthesis, other primary metabolite functions, Mildew Locus O genes (MLOs) and pathogenesis-related (PR) proteins. The expression of ten of these genes in both the MR-1 and Topmark cultivars was validated via qRT-PCR. Our results revealed both similar and differential patterns of gene expression between the two cultivars. This study may provide a new understanding of the molecular mechanisms of melon resistance to PM.

Sechium edule (Jacq.) Swartz is an important vegetable with both food and medicinal values. The complete chloroplast genome sequence of S. edule has been reported in this study. The total genome si...

The fungus Podosphaera xanthii causes powdery mildew, a disease that has serious negative effects on the production of melon (Cucumis melo L.) worldwide. In this study, an F2 melon population, derived from a cross between a resistant (MR-1) and a susceptible cultivar (Top Mark), was used for excavation of major powdery mildew resistance genes by bulked segregant analysis (BSA). The resistant and susceptible DNA bulks were constructed using 30 plants selected from the F2 population. Next-generation sequencing (NGS) was applied for the resequencing of the parental materials and two bulks. A total of 237.6 million paired reads were obtained that were used to calculate the Δsingle nucleotide polymorphisms (SNP)-index. A 0.6-Mb region on chromosome 12 was identified and further narrowed by 424 identified SNPs and quantitative trait locus (QTL) mapping in the F2 progeny with 25 screened polymorphic cleaved amplified polymorphic sequences (CAPS) markers. A major effective QTL named BPm12.1 was detected between the CAPS markers BSA12-LI3ECORI and BSA12-LI4HINFI which was tightly linked to the resistance gene for powdery mildew with genetic distances 0.02 cM and 0.28 cM. Seventeen candidate genes were identified, seven of which were predicted as candidate genes related to the resistance of melon to powdery mildew. The results of candidate gene analysis provided a potential target for further cloning and functional identification of the resistance gene in MR-1.

Citrullus mucosospermus (egusi watermelon) and Citrullus lanatus (dessert watermelon) are closely related subspecies. Variation in their seed coats underlies a key to understand the major differences in seeds between these two subspecies. Although the genetic model is understood, the candidate genes controlling the egusi seed trait in watermelon are unknown. In this study, a locus that controls the egusi seed trait, WEG6.1, which is located on chromosome 6 of the watermelon genome, was identified. A total of 195 polymorphic primers and 325 F2 plants were used to construct a primary mapping population and linkage maps. A large population of 758 F2 plants was used for fine mapping of the eg gene. The candidate gene was mapped to a 130.7 kb region on chromosome 6. Eleven genes were present within the corresponding region of the reference genome. All eleven candidate genes were sequenced, revealing that the CDS of the Cla007520 gene had a 3 bp deletion that resulted in the omission of one amino acid (serine) in C. mucosospermus. In addition, expression of the Cla007520 gene was highest at the early stage of seed formation, exhibiting a downward trend thereafter. The predicted protein of the Cla007520 gene belonged to the CPP protein family, and its amino acid constituents were similar to those of AtTSO1, which is associated with promoting integument cell proliferation in ovule development. This study revealed genes that facilitate the formation of egusi seed and might lead to a breakthrough in seed development and cultivated watermelon domestication.

Melons exhibit huge diversity of morphology and fruit quality traits, and these traits are still being investigated in worldwide breeding studies. Due to the fabulous advantages, single nucleotide polymorphisms (SNPs) based genetic markers are widely being used for dissection of putative QTLs/genes. In this study, we reported genetic linkage mapping and QTLs detection for ovary, fruit and seed traits of melon by using whole genome re-sequencing and newly developed SNPs based cleaved amplified polymorphism sequence (CAPS) markers. The re-sequencing analysis of two distinct melon lines exposed 14 million (96 %, 360 Mb) and 20 million (85.62 %, 319 Mb) re-sequenced reads, 404,083 candidate SNPs, and 6474 CAPS locus across the reference genome. An F2 mapping population comprising 148 individuals was genotyped with 163 SNP-CAPS markers for genetic linkage mapping which spanned total 1545.44 cM genetic distance and averaged 9.48 cM among whole genome adjacent markers. In total, 26 putative QTLs (6 major and 20 minor QTLs) were detected on differential chromosomes (1, 2, 3, 4, 6, 8, 9, and 12) and explained 1.83 %∼17.67 % phenotypic variations for 12 phenotypic traits with pleiotropic effects. Among these QTLs, 6 QTLs of ovary, 17 QTLs of fruit quality and 3 QTLs of seed traits were identified and many QTLs were clustered on chromosome 1, 9, and 12. A highly significant relationship and normal distribution frequency of phenotypes were also noticed in analysis of correlation and PCA. In crux, our detected QTL insights could be valuable and further investigated in unexplored botanical groups of melon by fine mapping and marker assisted selection (MAS).

A variety of melons are cultivated worldwide, and their specific biological properties make them an attractive model for molecular studies. This study aimed to investigate the single nucleotide polymorphisms (SNPs) from the mitochondrial, chloroplast, and nuclear genomes of seven melon accessions (Cucumis melo L.) to identify the phylogenetic relationships among melon cultivars with the Illumina HiSeq 2000 platform and bioinformatical analyses. The data showed that there were a total of 658 mitochondrial SNPs (207-295 in each), while there were 0-60 chloroplast SNPs among these seven melon cultivars, compared to the reference genome. Bioinformatical analysis showed that the mitochondrial tree topology was unable to separate the melon features, whereas the maximum parsimony/neighbor joining (MP/NJ) tree of the chloroplast SNPs could define melon features such as seed length, width, thickness, 100-seed weight, and type. SNPs of the nuclear genome were better than the mitochondrial and chloroplast SNPs in the identification of melon features. The data demonstrated the usefulness of mitochondrial, chloroplast, and nuclear SNPs in identification of phylogenetic associations among these seven melon cultivars.

Citrullus lanatus (watermelon) is an excellent daily source of dietary lycopene and β-carotene. To investigate the transcriptional regulation of carotenoid biosynthesis genes relative to lycopene and β-carotene accumulation in watermelon fruit, six watermelon accessions with different flesh colors were examined in this study: white-fleshed PI 459074, pale-yellow-fleshed ‘Cream of Saskatchewan’, light-pink-fleshed PI 482255, orange-yellow-fleshed ‘WM-Clr-1’, and red-fleshed ‘LSW177’ and ‘MSW28’. The expression patterns of eight genes (PSY1, PSY2, PDS, ZDS, CRTISO, LCYB, NCED1, and NCED7) involved in lycopene and β-carotene biosynthesis and biodegradation were analyzed. The results confirmed the accumulation of large quantities of lycopene in red-fleshed ‘LSW177’ and ‘MSW28’, reflecting the elevated expression of PSY1 and the low transcriptional expression of NCED1. The relative expression levels of NCED1 likely play an important role in the color development of the light-pink-fleshed PI 482255, whereas the reduced transcriptional expression of PSY1 and the increased expression of NCED1 appear to be the main factors contributing to the formation of white flesh in the fruit of PI 459074. Low transcriptional expression of PSY1 results in the pale-yellow flesh of the ‘Cream of Saskatchewan’ fruit.

Cleaved amplified polymorphic sequence (CAPS) markers are useful tools for detecting single nucleotide polymorphisms (SNPs). This study detected and converted SNP sites into CAPS markers based on high-throughput re-sequencing data in watermelon, for linkage map construction and quantitative trait locus (QTL) analysis. Two inbred lines, Cream of Saskatchewan (COS) and LSW-177 had been re-sequenced and analyzed by Perl self-compiled script for CAPS marker development. 88.7% and 78.5% of the assembled sequences of the two parental materials could map to the reference watermelon genome, respectively. Comparative assembled genome data analysis provided 225,693 and 19,268 SNPs and indels between the two materials. 532 pairs of CAPS markers were designed with 16 restriction enzymes, among which 271 pairs of primers gave distinct bands of the expected length and polymorphic bands, via PCR and enzyme digestion, with a polymorphic rate of 50.94%. Using the new CAPS markers, an initial CAPS-based genetic linkage map was constructed with the F2 population, spanning 1836.51 cM with 11 linkage groups and 301 markers. 12 QTLs were detected related to fruit flesh color, length, width, shape index, and brix content. These newly CAPS markers will be a valuable resource for breeding programs and genetic studies of watermelon.

Citrullus colocynthis L. is one of the worldwide famous traditionally medicinal plants and widely applied in watermelon breeding for its multiple resistances. The complete nucleotide sequence of desert watermelon (Citrullus colocythis L.) chloroplast genome has been determined in this study. The genome was composed of 157,147 bp containing a pair of inverted repeats (IRs) of 26,149 bp, which was separated by a large single-copy region of 86,851 bp and a small single-copy region of 17,998 bp. A total of 123 genes were predicted including 86 protein-coding genes, eight rRNA genes and 29 tRNA genes. Phylogenetic analysis revealed that C. colocynthis were closely related to other two species in the genus Citrullus. The complete chloroplast genome of C. colocynthis would provide some significant information for Cucurbitaceae evolutionary and genomic studies.

Genetic basis of melon morphology has been well-studied over the past few years. Single nucleotide polymorphisms (SNPs) based genetic markers are widely being used for QTLs and gene(s) detection. In this study, we reported genetic linkage mapping and QTLs detection of ovary-related traits in melon using whole genome re-sequencing and newly developed SNPs based cleaved amplified polymorphism sequence (CAPS) markers. Re-sequencing analysis exhibited that two distinct parental lines of melon covered 97.48% and 96.02% of reference genome, and total 302,684 major SNPs yielded 3519 CAPS loci. An F2 mapping population of 156 individuals was genotyped for genetic linkage mapping with novel 149 SNP-CAPS markers, and spanning total 4072 cM distance and 27.32 cM in average b/w adjacent markers on 12 chromosomes. In total, 7 detected QTLs on different chromosomes (1, 5, 7, 8 and 11) explained 4.10%–17.78% phenotypic variations for 5 ovary-related traits and QTL region for ovary hairs density on chromosome 7 is mapped first time in this study. The detected QTLs regions revealed total 134 genes at different narrow genomic regions (365.28 (OVL), 343.89 (OVW), 23.47 (OVS), 107.05 (OVHD) and 597.09 (OVHL)) in kbp b/w adjacent CAPS markers on various chromosomes. Normal distribution frequency and highly significant correlation was also noticed in phenotypic analysis. Our results provide the theoretical basis of multiple QTL insights for ovary-related traits in melon and could be further investigated in unexplored botanical groups of melon by cloning and functional characterization in future studies.

Melon (Cucumis melo. L.) is an economically important vegetable crop and a model species for the study of sex determination. Although two master genes, G (for gynoecy) and A (andromonoecy) have been cloned, the process of sex expression in melon is complex, and the associated mechanisms and gene network are not well understood. In this study, RNA sequencing was conducted among bulk segregants of four melon plant sex types, namely monoecious (AAGG), gynoecious (AAgg), hermaphrodite (aagg), and andromonoecious (aaGG). About 105 million reads were generated from the melon transcriptome using Illumina HiSeq 2000 sequencing. In total, 76,260 unigenes were generated and mapped to 11,805 annotated proteins in the assembled melon genome. Most of the genes encode proteins related to plant hormone metabolism. Others are related to flora development, including tasselseeds and male sterility genes, which were detected in the phytohormone pathway. Comparisons of paired segregants (AAGG:AAgg, AAGG:aaGG, aagg:AAgg and aaGG:aagg) revealed different gene expression profiles (745, 1342, 858, and 571 genes, respectively). The most highly represented was the serine/threonine protein kinase pathway. In this pathway, ethylene, abscisic acid (ABA), auxin/indole-3-acetic acid (IAA), and aminocyclopropanecarboxylate (ACC) oxidase were involved in different sex type expressions, suggesting that they might have important roles in melon sex determination. Fifteen selected genes were validated via reverse transcription quantitative real-time polymerase chain reaction, and their expression patterns were similar for the two methods. Overall, the melon transcriptome sequences revealed novel gene expression profiles and offer important clues for further study of the molecular mechanism of sex differentiation.

Lycopene content is an important factor for determining watermelon fruit quality. However, the low DNA polymorphism among cultivated watermelon (Citrullus lanatus) has hindered the ability to establish high quality genetic maps and study the quantitative trait loci (QTL) controlling the lycopene content trait. In this study, we successfully constructed a genetic map of watermelon to determine lycopene content and other horticultural fruit traits using a F2 population developed from a cross between the two lines of watermelon LSW-177 and Cream of Saskatchewan. The genetic map contained 16 linkage groups covering a total length of 2,039.5 cM, which included 37 SSRs (Simple Sequence Repeat) and 107 CAPSs (Cleaved Amplified Polymorphic Sequences), with all of the CAPS markers developed from high-throughput re-sequencing of data from this study. Three CAPS markers (WII04E07-33,WII04E07-37,WII04E07-40) caused the F2 population to perfectly co-segregate for each F2 population plants. We also obtained 12 QTLs for all of the traits measured. Only one QTL (LCYB4.1) was detected with a high value of trait variation (83.50 %) that related to lycopene content and mapped on Chromosome 4 between CAPS markers WII04E07-33 and WII04E07-40, which could nearly account for all of the differences in lycopene content between the two parental strains. In this study, we highlighted 2,458 CAPS loci that were suitable for primer design with a polymorphism of 48.9 %, which is approximately a 12-fold increase from previous studies. The present map and QTLs will facilitate future studies on determining lycopene content related genes and cloning watermelon genes, while also providing for useful markers for breeding for lycopene content.

Melon is an important agricultural and economic vegetable crop worldwide. The genetic male sterility mutant (ms-5) has a recessive nuclear gene that controls the male sterility germplasm. Male sterility could reduce the cost of F1 seed production in melon, but heterozygous fertile plants should be removed before pollination. In this study, bulked segregant analysis combined with specific length amplified fragment sequencing was applied to map the single nuclear male sterility recessive gene. A 30-kb candidate region on chromosome 9 located on scaffold 000048 and spanning 2,522,791 to 2,555,104 bp was identified and further confirmed by cleavage amplified polymorphic sequence markers based on parental line resequencing data and classical mapping of 252 F2 individuals. Gene prediction indicated that six annotated genes are present in the 30-kb candidate region. Quantitative RT-PCR revealed significant differences in the expression level of the LOC103498166 ABORTED MICROSPORES (AMS) gene in male-sterile lines (ms-5) and male-fertile (HM1-1) lines during the 2-mm (tetrad) and 5-mm (the first pollen mitosis) periods, and negative regulation of the AMS candidate gene transcription factor was also detected. Sequencing and cluster analysis of the AMS transcription factor revealed five single-nucleotide polymorphisms between the parental lines. The data presented herein suggest that the AMS transcription factor is a possible candidate gene for single nuclear male sterility in melon. The results of this study will help breeders to identify male-sterile and -fertile plants at seeding as marker-assisted selection methods, which would reduce the cost of seed production and improve the use of male-sterile lines in melon.

Simple sequence repeats (SSRs) are one of the most informative and widely used molecular markers in plant research. The melon draft genome has provided a powerful tool for SSR marker development in this species in which there are still not enough SSR markers. We therefore developed genome-wide SSR markers from melon, which were used for genetic diversity analysis in melon accessions and comparative mapping with cucumber and watermelon. A total of 44,265 microsatellites from the melon genome were characterized, of which 28,570 SSR markers were developed. In silico PCR analysis with these SSR markers identified 4002 and 1085 with one amplicon in cucumber and watermelon genome, respectively. With these cross-species transferable melon SSR markers, the chromosome synteny between melon and cucumber as well as watermelon was established, which revealed complicated mosaic patterns of syntenic blocks among them. We experimentally validated 384 SSR markers, from which 42 highly informative SSR ones were selected for genetic diversity and population structure analysis among 118 melon accessions. The large number of melon SSR markers developed in this study provides a valuable resource for genetic linkage map construction, molecular mapping, and marker-assisted selection (MAS) in melon. Furthermore, the cross-species transferable SSR markers could also be useful in various molecular marker-related studies in other closely related species in Cucurbitaceae family in which draft genomes are not yet available.

Cucurbita pepo is an important economic plant cultivated widely in the world. The complete chloroplast genome sequence of C. pepo is reported here. The genome is 157,343 bp in length and ex...

The aim of the present study was to develop a transformation system that may be useful for introducing agronomically and biotechnologically relevant traits into melon. The production of transplanted melon with maternal inheritance of the transgene could solve problems related to outcrossing between genetically modified crops and conventional crops or their wild relatives. By analyzing the main influencing factors systematically, the pollination time was ascertained and the pollen-tube pathway genetic transformation system was optimized. A screening system for resistant seeds from the T1 generation was established. The transformed seedlings were grown under standard field conditions and selected using a polymerase chain reaction-based analysis. The resistant plants were detected at a rate of 5%. These results indicate that enhanced production hastens the initiation of bisexual flowers, development of mature bisexual flowers, and fruit set in melon. We have established a melon transformation system based on the pollen-tube method.

Cucumber, Cucumis sativus L. is the only taxon with 2n = 2x = 14 chromosomes in the genus Cucumis. It consists of two cross-compatible botanical varieties: the cultivated C. sativus var. sativus and the wild C. sativus var. hardwickii. There is no consensus on the evolutionary relationship between the two taxa. Whole-genome sequencing of the cucumber genome provides a new opportunity to advance our understanding of chromosome evolution and the domestication history of cucumber. In this study, a high-density genetic map for cultivated cucumber was developed that contained 735 marker loci in seven linkage groups spanning 707.8 cM. Integration of genetic and physical maps resulted in a chromosome-level draft genome assembly comprising 193 Mbp, or 53% of the 367 Mbp cucumber genome. Strategically selected markers from the genetic map and draft genome assembly were employed to screen for fosmid clones for use as probes in comparative fluorescence in situ hybridization analysis of pachytene chromosomes to investigate genetic differentiation between wild and cultivated cucumbers. Significant differences in the amount and distribution of heterochromatins, as well as chromosomal rearrangements, were uncovered between the two taxa. In particular, six inversions, five paracentric and one pericentric, were revealed in chromosomes 4, 5 and 7. Comparison of the order of fosmid loci on chromosome 7 of cultivated and wild cucumbers, and the syntenic melon chromosome I suggested that the paracentric inversion in this chromosome occurred during domestication of cucumber. The results support the sub-species status of these two cucumber taxa, and suggest that C. sativus var. hardwickii is the progenitor of cultivated cucumber.

In this study, molecular markers were designed based on the sex determination genes ACS7 (A) and WIP1 (G) and the domain in the Fusarium oxysporum-resistant gene Fom-2 (F) in order to achieve selection of F. oxysporum-resistant gynoecious melon plants. Markers of A and F are cleaved amplified polymorphic sequences that distinguish alleles according to restriction analysis. Twenty F1 and 1863 F2 plants derived from the crosses between the gynoecious line WI998 and the Fusarium wilt-resistant line MR-1 were genotyped based on the markers. The results showed that the polymerase chain reaction and enzyme digestion results could be effectively used to identify plants with the AAggFF genotype in F2 populations. In the F2 population, 35 gynoecious wilt-resistant plants were selected by marker-assisted selection and were confirmed by disease infection assays, demonstrating that these markers can be used in breeding to select F. oxysporum-resistant gynoecious melon plants.

The inheritance of sex determination genes in melon was studied using the populations of F 1 , F 2 , BC 1 P 1 and BC 1 P 2 derived from two parental lines, the gynoecious WI998 and andromonoecious TopMark. These populations were segregating at two loci for sex determination: the a gene locus for andromonoecious and the m gene locus for gynoecious sex expression. It was found that different phenotypes corresponded to different genotypes in sex expression: A_G_ = monoecious; A_gg = gynomonecious; aaG_ = andromonoecious; aagg = hemaphrodite; and A_ggmm = gynoecious. A preliminary genetic mapping of these two gene loci were conducted with microsatellite markers in the F 2 population with 298 individuals. Fifty-five polymorphic SSR markers were identified and applied for linkage analysis to construct the genetic map which spanned 666.7 cM with a mean marker interval of 17.09 cM. SSR marker MU5549-1 linked to gene a with the distance at 13.5 cM and SSR marker MU14723-2 linked to gene m with the distance at 11.6 cM.

Genetic relatedness and phenotype are important factors that govern the expression of heterosis in hybrid progeny of many cross-pollinating plant species. Since this relationship is important but not well understood in melon (Cucumis melo L.), one monoecious and two andromonoecious melon lines of diverse Chinese [Peoples Republic of China (PRC)] origin were crossed to the andromonoecious U.S. Western Shipping market type ‘Top Mark’ (TM) and the andromonoecious, highly branched line H-16 to determine parental combining ability and heterosis for five yield component traits in three test environments [open-field (USA), and energy-saving (PRC) and plastic greenhouses (PRC)]. Random amplified polymorphic DNA (RAPD)- and simple sequence repeat (SSR)-based genetic distances (GD) among and between parents (5) and their hybrids (6) were calculated and compared to phenotypic trait values. These germplasms were evaluated for lateral branch number (LBN), days to 50% flower (DF), fruit number and weight per plant, and fruit length:diameter (L:D) ratio in each of three test environments. General combining ability was significant for all characters, except for L:D in all locations, and LBN and DF in the plastic greenhouse environment. Both descriptors of difference (genetic marker and phenotype) were discriminatory, and provided similar assessments of relationships among parents and hybrids. Although dramatic performance differences were detected between parents and among F1 hybrid progeny, a strong relationship between GD and heterotic effects was not consistently detected.