Clpf encodes pentatricopeptide repeat protein (PPR5) and regulates pink flesh color in watermelon (Citrullus lanatus L.).

KEY MESSAGE: The gene controlling pink flesh in watermelon was finely mapped to a 55.26-kb region on chromosome 6. The prime candidate gene, Cla97C06G122120 (ClPPR5), was identified through forward genetics. Carotenoids offer numerous health benefits; while, they cannot be synthesized by the human body. Watermelon stands out as one of the richest sources of carotenoids. In this study, genetic generations derived from parental lines W15-059 (red flesh) and JQ13-3 (pink flesh) revealed the presence of the recessive gene Clpf responsible for the pink flesh (pf) trait in watermelon. Comparative analysis of pigment components and microstructure indicated that the disparity in flesh color between the parental lines primarily stemmed from variations in lycopene content, as well as differences in chromoplast number and size. Subsequent bulk segregant analysis (BSA-seq) and genetic mapping successfully narrowed down the Clpf locus to a 55.26-kb region on chromosome 6, harboring two candidate genes. Through sequence comparison and gene expression analysis, Cla97C06G122120 (annotated as a pentatricopeptide repeat, PPR) was predicted as the prime candidate gene related to pink flesh trait. To further investigate the role of the PPR gene, its homologous gene in tomato was silenced using a virus-induced system. The resulting silenced fruit lines displayed diminished carotenoid accumulation compared with the wild-type, indicating the potential regulatory function of the PPR gene in pigment accumulation. This study significantly contributes to our understanding of the forward genetics underlying watermelon flesh traits, particularly in relation to carotenoid accumulation. The findings lay essential groundwork for elucidating mechanisms governing pigment synthesis and deposition in watermelon flesh, thereby providing valuable insights for future breeding strategies aimed at enhancing fruit quality and nutritional value.

Optimized combination methods for exploring and verifying disease-resistant transcription factors in melon.

A large amount of omics data and number of bioinformatics tools has been produced. However, the methods for further exploring omics data are simple, in particular, to mine key regulatory genes, which are a priority concern in biological systems, and most of the specific functions are still unknown. First, raw data of two genotypes of melon (susceptible and resistant) were obtained by transcriptome analysis. Second, 391 transcription factors (TFs) were identified from the plant transcription factor database and cucurbit genomics database. Then, functional enrichment analysis indicated that these genes were mainly annotated in the process of transcription regulation. Third, 243 and 230 module-specific TFs were screened by weighted gene coexpression network analysis and short time series expression miner, respectively. Several TF genes, such as WRKYs and bHLHs, were regarded as key regulatory genes according to the values of significantly different modules. The coexpression network showed that these TF genes were significant correlated with resistance (R) genes, such as DRP2, RGA3, DRP1 and NB-ARC. Fourth, cis-acting element analysis illustrated that these R genes may bind to WRKY and bHLH. Finally, the expression of WRKY genes was verified by quantitative reverse transcription PCR (RT-qPCR). Phylogenetic analysis was carried out to further confirm that these TFs may play a critical role in Curcurbitaceae disease resistance. This study provides a new optimized combination strategy to explore the functions of TFs in a wide spectrum of biological processes. This strategy may also effectively predict potential relationships in the interactions of essential genes.

Integrated analysis of biparental and natural populations reveals CRIB domain-containing protein underlying seed coat crack trait in watermelon.

KEY MESSAGE: The scc locus of the watermelon seed coat crack trait was fine mapped on chromosome 3. Cla97C03G056110 (annotated as CRIB domain-containing protein) was regarded as the most likely candidate gene Seed coat crack (scc) is a special characteristic of watermelon compared with other cucurbit crops. However, information regarding the genetic basis of this trait is limited. We conducted a genetic analysis of six generations derived from PI 192938 (scc) and Cream of Saskatchewan (COS) (non-scc) parental lines and found that the scc trait was regulated by a single recessive gene through two years. Bulk segregant analysis sequencing (BSA-seq) and initial mapping placed the scc locus into an 808.8 kb region on chromosome 3. Evaluation of another 1152 F2 plants narrowed the scc locus to a 277.11 kb region containing 37 candidate genes. Due to the lack of molecular markers in the fine-mapping interval, we extracted the genome sequence variations in this 277.11 kb region with in silico BSA among seventeen re-sequenced lines (6 scc and 11 non-scc) and finally delimited the scc locus to an 8.34 kb region with only one candidate gene Cla97C03G056110 (CRIB domain-containing protein). Three single nucleotide polymorphism loci in the promoter region of Cla97C03G056110 altered cis-acting elements that were highly correlated with the nature watermelon panel. The expression of Cla97C03G056110 in seed coat tissue was higher in non-scc than in scc lines and was specifically expressed in seed coat compared with fruit flesh.

A recessive gene Cmpmr2F confers powdery mildew resistance in melon (Cucumis melo L.).

KEY MESSAGE: Identified a recessive gene (Cmpmr2F) associated with resistance to infection by the powdery mildew causing agent Podosphaera xanthii race 2F. Powdery mildew (PM) is one of the most destructive fungal diseases of melon, which significantly reduces the crop yield and quality. Multiple studies are being performed for in-depth genetic understandings of PM-susceptibility or -resistance mechanisms in melon plants, but the holistic knowledge of the precise genetic basis of PM-resistance is unexplored. In this study, we characterized the recessive gene "Cmpmr2F" and found its association with resistance against the PM causative agent "Podosphaera xanthii race 2F." Fine genetic mapping revealed the major-effect region of a 26.25-kb interval on chromosome 12, which harbored the Cmpmr2F gene corresponding to the MELO3C002403, encoding allantoate amidohydrolase. The functional gene annotation, expression pattern, and sequence alignment analyses were carried out using two contrast parent lines of melon "X055" PM-susceptible and "PI 124112" PM-resistant. Further, gene silencing of Cmpmr2F using virus-induced gene silencing (VIGS) significantly increased PM-resistance in the susceptible plant. In contrast to the previously reported studies, we identified that Cmpmr2F-silenced plants showed no impairment in growth due to less apparent negative effects in silenced melon plants. So, it is believed that the Cmpmr2F gene has great potential for further breeding studies to increase the P. xanthii race 2F resistance in melon. In short, our study provides new genetic resources and a solid foundation for further functional analysis of PM-resistance genes in melon, as well as powerful molecular markers for marker-assisted breeding aimed at developing new melon varieties resistant to PM infection.

Molecular Mapping of Putative Genomic Regions Controlling Fruit and Seed Morphology of Watermelon.

The genetic regulatory basis of qualitative and quantitative phenotypes of watermelon is being investigated in different types of molecular and genetic breeding studies around the world. In this study, biparental F2 mapping populations were developed over two experimental years, and the collected datasets of fruit and seed traits exhibited highly significant correlations. Whole-genome resequencing of comparative parental lines was performed and detected single nucleotide polymorphism (SNP) loci were converted into cleaved amplified polymorphic sequence (CAPS) markers. The screened polymorphic markers were genotyped in segregating populations and two genetic linkage maps were constructed, which covered a total of 2834.28 and 2721.45 centimorgan (cM) genetic lengths, respectively. A total of 22 quantitative trait loci (QTLs) for seven phenotypic traits were mapped; among them, five stable and major-effect QTLs (PC-8-1, SL-9-1, SWi-9-1, SSi-9-1, and SW-6-1) and four minor-effect QTLs (PC-2-1 and PC-2-2; PT-2-1 and PT-2-2; SL-6-1 and SSi-6-2; and SWi-6-1 and SWi-6-2) were observed with 3.77-38.98% PVE. The adjacent QTL markers showed a good fit marker-trait association, and a significant allele-specific contribution was also noticed for genetic inheritance of traits. Further, a total of four candidate genes (Cla97C09G179150, Cla97C09G179350, Cla97C09G180040, and Cla97C09G180100) were spotted in the stable colocalized QTLs of seed size linked traits (SL-9-1 and SWi-9-1) that showed non-synonymous type mutations. The gene expression trends indicated that the seed morphology had been formed in the early developmental stage and showed the genetic regulation of seed shape formation. Hence, we think that our identified QTLs and genes would provide powerful genetic insights for marker-assisted breeding aimed at improving the quality traits of watermelon.

CmPMRl and CmPMrs are responsible for resistance to powdery mildew caused by Podosphaera xanthii race 1 in Melon.

KEY MESSAGE: 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 (P1; resistant to PM) and M4-7 (P2; susceptible to PM) accessions to assess the heritability of PM (race 1) resistance in these melon plants. PM resistance in MR-1 leaves was linked to a dominant gene (CmPMRl), whereas stem resistance was under the control of a recessive gene (CmPMrs), with the dominant gene having an epistatic effect on the recessive gene. The CmPMRl gene was mapped to a 50 Kb interval on chromosome 12, while CmPMrs was mapped to an 89 Kb interval on chromosome 10. The CmPMRl candidate gene MELO3C002441 and the CmPMrs candidate gene MELO3C012438 were identified through sequence alignment, functional annotation, and expression pattern analyzes of all genes within these respective intervals. MELO3C002441 and MELO3C012438 were both localized to the cellular membrane and were contained conserved NPR gene-like and MLO domains, respectively, which were linked to PM resistance. In summary, we identified patterns of PM resistance in the disease-resistant MR-1 melon cultivar and identified two putative genes linked to resistance. Our results offer new genetic resources and markers to guide future marker-assisted breeding for PM resistance in melon.

Nucleotide variation in the phytoene synthase (ClPsy1) gene contributes to golden flesh in watermelon (Citrullus lanatus L.).

KEY MESSAGE: A gene controlling golden flesh trait in watermelon was discovered and fine mapped to a 39.08 Kb region on chromosome 1 through a forward genetic strategy, and Cla97C01G008760 (annotated as phytoene synthase protein, ClPsy1 ) was recognized as the most likely candidate gene. Vitamin A deficiency is a worldwide public nutrition problem, and ß-carotene is the precursor for vitamin A synthesis. Watermelon with golden flesh (gf, which occurs due to an accumulated abundance of ß-carotene) is an important germplasm resource. In this study, a genetic analysis of segregated gf gene populations indicated that gf was controlled by a single recessive gene. BSA-seq (Bulked segregation analysis) 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 including over 1000 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 the gf trait. Nonsynonymous SNP mutations in the first exon of ClPsy1 between parental lines co-segregated with the gf trait only among individuals in the genetic population and 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 exhibited expression trends similar to that of ClPsy1 in the parental lines and may regulate the ClPsy1 expression. Further 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 watermelon ß-carotene accumulation.

Biparental genetic mapping reveals that CmCLAVATA3 (CmCLV3) is responsible for the variation in carpel number in melon (Cucumis melo L.).

KEY MESSAGES: 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 F2 population containing 1026 individuals further narrowed the locus to an 83.98 kb region harboring five annotated genes. Gene structure alignment between the parental lines revealed MELO3C035640.2 (annotated as CLAVATA3, CmCLV3) as the best candidate gene for the CN trait. CmCLV3 was more highly expressed in 3- than 5-CN lines and specifically expressed in terminal buds rather than in young leaves, hypocotyls, and roots. The CmCLV3 coding region was cloned from eight 3- or 5-CN melon accessions, and a nonsynonymous SNP site was highly correlated with CN variation. This SNP site was also related to CN variations among 40 melon lines according to their resequencing data, causing a helix alteration in the CmCLV3 protein. Promoter region sequence alignment and activity analysis showed that, unlike in cucumber and tomato, CmCLV3 promoter variation and activity were not the main reasons for CN alteration. Overall, this study provides a genetic resource for melon fruit development research and molecular breeding tools for melon CN improvement.

Mapping of genetic loci controlling fruit linked morphological traits of melon using developed CAPS markers.

BACKGROUND: 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. METHODS AND RESULTS: 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-F2 derived plant populations were genotyped for linkage mapping and composite interval mapping based QTL mapping exposed 6 genetic loci, positioned over distinct chromosomes (02, 04, 08, 09, and 12) between the flanking intervals of CAPS markers, which explained an unlinked polygenic architecture in genome. Three minor QTLs of fruit weight (FWt2.1, FWt4.1, FWt9.1), one major QTL of fruit firmness (FrFir8.1), one major QTL of fruit length (FL12.1), and one major QTL of fruit shape (FS12.1) were determined and collectively explained the phenotypic variance from 5.64 to 15.64%. Fruit phenotypic correlation exhibited the significant relationship and principal component analysis also identified the potential variability. Multiple sequence alignments also indicated the significant base-mutations in the detected genetic loci, respectively. CONCLUSION: In 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.

Genome-Wide Analysis of the Peroxidase Gene Family and Verification of Lignin Synthesis-Related Genes in Watermelon.

Watermelon (Citrullus lanatus) is an important horticultural crop worldwide, but peel cracking caused by peel hardness severely decreases its quality. Lignification is one of the important functions of class III peroxidase (PRX), and its accumulation in the plant cell wall leads to cell thickening and wood hardening. For in-depth physiological and genetical understanding, we studied the relationship between peel hardness and lignin accumulation and the role of PRXs affecting peel lignin biosynthesis using genome-wide bioinformatics analysis. The obtained results showed that lignin accumulation gradually increased to form the peel stone cell structure, and tissue lignification led to peel hardness. A total of 79 ClPRXs (class III) were identified using bioinformatics analysis, which were widely distributed on 11 chromosomes. The constructed phylogenetics indicated that ClPRXs were divided into seven groups and eleven subclasses, and gene members of each group had highly conserved intron structures. Repeated pattern analysis showed that deletion and replication events occurred during the process of ClPRX amplification. However, in the whole-protein sequence alignment analysis, high homology was not observed, although all contained four conserved functional sites. Repeated pattern analysis showed that deletion and replication events occurred during ClPRXs' amplification process. The prediction of the promoter cis-acting element and qRT-PCR analysis in four tissues (leaf, petiole, stem, and peel) showed different expression patterns for tissue specificity, abiotic stress, and hormone response by providing a genetic basis of the ClPRX gene family involved in a variety of physiological processes in plants. To our knowledge, we for the first time report the key roles of two ClPRXs in watermelon peel lignin synthesis. In conclusion, the extensive data collected in this study can be used for additional functional analysis of ClPRXs in watermelon growth and development and hormone and abiotic stress response.

Comparative Transcriptome Analysis Identified Key Pathways and Genes Regulating Differentiated Stigma Color in Melon (Cucumis melo L.).

Stigma color is an important morphological trait in many flowering plants. Visual observations in different field experiments have shown that a green stigma in melons is more attractive to natural pollinators than a yellow one. In the current study, we evaluated the characterization of two contrasted melon lines (MR-1 with a green stigma and M4-7 with a yellow stigma). Endogenous quantification showed that the chlorophyll and carotenoid content in the MR-1 stigmas was higher compared to the M4-7 stigmas. The primary differences in the chloroplast ultrastructure at different developmental stages depicted that the stigmas of both melon lines were mainly enriched with granum, plastoglobulus, and starch grains. Further, comparative transcriptomic analysis was performed to identify the candidate pathways and genes regulating melon stigma color during key developmental stages (S1-S3). The obtained results indicated similar biological processes involved in the three stages, but major differences were observed in light reactions and chloroplast pathways. The weighted gene co-expression network analysis (WGCNA) of differentially expressed genes (DEGs) uncovered a "black" network module (655 out of 5302 genes), mainly corresponding to light reactions, light harvesting, the chlorophyll metabolic process, and the chlorophyll biosynthetic process, and exhibited a significant contribution to stigma color. Overall, the expression of five key genes of the chlorophyll synthesis pathway-CAO (MELO03C010624), CHLH (MELO03C007233), CRD (MELO03C026802), HEMA (MELO03C011113), POR (MELO03C016714)-were checked at different stages of stigma development in both melon lines using quantitative real time polymerase chain reaction (qRT-PCR). The results exhibited that the expression of these genes gradually increased during the stigma development of the MR-1 line but decreased in the M4-7 line at S2. In addition, the expression trends in different stages were the same as RNA-seq, indicating data accuracy. To sum up, our research reveals an in-depth molecular mechanism of stigma coloration and suggests that chlorophyll and related biological activity play an important role in differentiating melon stigma color.

Genome-wide identification, characterization and expression analysis of the TLP gene family in melon (Cucumis melo L.).

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.

Resequencing of 297 melon accessions reveals the genomic history of improvement and loci related to fruit traits in melon.

Domestication and improvement are two important stages in crop evolution. Melon (Cucumis melo L.) is an important vegetable crop with wide phenotypic diversity in many horticultural traits, especially fruit size, flesh thickness and aroma, which are likely the results of long-term extensive selection during its evolution. However, selective signals in domestication and improvement stages for these remarkable variations remain unclear. We resequenced 297 wild, landrace and improved melon accessions and obtained 2 045 412 high-quality SNPs. Population structure and genetic diversity analyses revealed independent and two-step selections in two subspecies of melon: ssp. melo and ssp. agrestis during melon breeding. We detected 233 (~18.35 Mbp) and 159 (~17.71 Mbp) novel potential selective signals during the improvement stage in ssp. agrestis and spp. melo, respectively. Two alcohol acyltransferase genes (CmAATs) unique to the melon genome compared with other cucurbit crops may have undergone stronger selection in ssp. agrestis for the characteristic aroma as compared with other cucurbits. Genome-wide association analysis identified eight fruit size and seven flesh thickness signals overlapping with selective sweeps. Compared with thin-skinned ssp. agrestis, thick-skinned ssp. melo has undergone a stronger selection for thicker flesh. In most melon accessions, CmCLV3 has pleiotropic effects on carpel number and fruit shape. Findings from this study provide novel insights into melon crop evolution, and new tools to advance melon breeding.

Genetic architecture of fruit size and shape variation in cucurbits: a comparative perspective.

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.

Comparative transcriptome analysis of two contrasting watermelon genotypes during fruit development and ripening.

BACKGROUND: Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] is an economically important crop with an attractive ripe fruit that has colorful flesh. Fruit ripening is a complex, genetically programmed process. RESULTS: In this study, a comparative transcriptome analysis was performed to identify the regulators and pathways that are involved in the fruit ripening of pale-yellow-flesh cultivated watermelon (COS) and red-flesh cultivated watermelon (LSW177). We first identified 797 novel genes to extend the available reference gene set. Second, 3958 genes in COS and 3503 genes in LSW177 showed at least two-fold variation in expression, and a large number of these differentially expressed genes (DEGs) during fruit ripening were related to carotenoid biosynthesis, plant hormone pathways, and sugar and cell wall metabolism. Third, we noted a correlation between ripening-associated transcripts and metabolites and the key function of these metabolic pathways during fruit ripening. CONCLUSION: The results revealed several ripening-associated actions and provide novel insights into the molecular mechanisms underlying the regulation of watermelon fruit ripening.

Comparative analysis of single nucleotide polymorphisms in the nuclear, chloroplast, and mitochondrial genomes in identification of phylogenetic association among seven melon (Cucumis melo L.) cultivars.

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.

Development of cleaved amplified polymorphic sequence markers and a CAPS-based genetic linkage map in watermelon (Citrullus lanatus [Thunb.] Matsum. and Nakai) constructed using whole-genome re-sequencing data.

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.

Next-generation sequencing, FISH mapping and synteny-based modeling reveal mechanisms of decreasing dysploidy in Cucumis.

In the large Cucurbitaceae genus Cucumis, cucumber (C. sativus) is the only species with 2n = 2x = 14 chromosomes. The majority of the remaining species, including melon (C. melo) and the sister species of cucumber, C. hystrix, have 2n = 2x = 24 chromosomes, implying a reduction from n = 12 to n = 7. To understand the underlying mechanisms, we investigated chromosome synteny among cucumber, C. hystrix and melon using integrated and complementary approaches. We identified 14 inversions and a C. hystrix lineage-specific reciprocal inversion between C. hystrix and melon. The results reveal the location and orientation of 53 C. hystrix syntenic blocks on the seven cucumber chromosomes, and allow us to infer at least 59 chromosome rearrangement events that led to the seven cucumber chromosomes, including five fusions, four translocations, and 50 inversions. The 12 inferred chromosomes (AK1-AK12) of an ancestor similar to melon and C. hystrix had strikingly different evolutionary fates, with cucumber chromosome C1 apparently resulting from insertion of chromosome AK12 into the centromeric region of translocated AK2/AK8, cucumber chromosome C3 originating from a Robertsonian-like translocation between AK4 and AK6, and cucumber chromosome C5 originating from fusion of AK9 and AK10. Chromosomes C2, C4 and C6 were the result of complex reshuffling of syntenic blocks from three (AK3, AK5 and AK11), three (AK5, AK7 and AK8) and five (AK2, AK3, AK5, AK8 and AK11) ancestral chromosomes, respectively, through 33 fusion, translocation and inversion events. Previous results (Huang, S., Li, R., Zhang, Z. et al., , Nat. Genet. 41, 1275-1281; Li, D., Cuevas, H.E., Yang, L., Li, Y., Garcia-Mas, J., Zalapa, J., Staub, J.E., Luan, F., Reddy, U., He, X., Gong, Z., Weng, Y. 2011a, BMC Genomics, 12, 396) showing that cucumber C7 stayed largely intact during the entire evolution of Cucumis are supported. Results from this study allow a fine-scale understanding of the mechanisms of dysploid chromosome reduction that has not been achieved previously.

Identification of Chromosomal Regions and Candidate Genes for Round leaf Locus in Cucumis melo L.

Leaf morphology plays a crucial role in plant classification and provides a significant model for studying plant diversity while directly impacting photosynthetic efficiency. In the case of melons, leaf shape not only influences production and classification but also represents a key genetic trait that requires further exploration. In this study, we utilized forward genetics to pinpoint a recessive locus, dubbed Cmrl (Round leaf), which is responsible for regulating melon leaf shape. Through bulked segregant analysis sequencing and extensive evaluation of a two-year F2 population, we successfully mapped the Cmrl locus to a 537.07 kb region on chromosome 8 of the melon genome. Subsequent genetic fine-mapping efforts, leveraging a larger F2 population encompassing 1322 plants and incorporating F2:3 phenotypic data, further refined the locus to an 80.27 kb interval housing five candidate genes. Promoter analysis and coding sequence cloning confirmed that one of these candidates, MELO3C019152.2 (Cmppr encoding a pentatricopeptide repeat-containing family protein, Cmppr), stands out as a strong candidate gene for the Cmrl locus. Notably, comparisons of Cmrl expressions across various stages of leaf development and different leaf regions suggest a pivotal role of Cmrl in the morphogenesis of melon leaves.