Mapping and validation of Fusarium wilt race 2 resistance QTL from Citrullus amarus line USVL246-FR2.

KEY MESSAGE: Fon race 2 resistant QTLs were identified on chromosomes 8 and 9. Families homozygous for resistance alleles at a haplotype of three KASP markers had 42% lower disease severity than those with susceptible alleles in an independent, interspecific validation population confirming their utility for introgression of Fusarium wilt resistance. Fusarium oxysporum f. sp. niveum (Fon) race 2 causes Fusarium wilt in watermelon and threatens watermelon production worldwide. Chemical management options are not effective, and no resistant edible watermelon cultivars have been released. Implementation of marker-assisted selection to develop resistant cultivars requires identifying sources of resistance and the underlying quantitative trait loci (QTL), developing molecular markers associated with the QTL, and validating marker-phenotype associations with an independent population. An intraspecific Citrullus amarus recombinant inbred line population from a cross of resistant USVL246-FR2 and susceptible USVL114 was used for mapping Fon race 2 resistance QTL. KASP markers were developed (N = 51) for the major QTL on chromosome 9 and minor QTL on chromosomes 1, 6, and 8. An interspecific F2:3 population was developed from resistance donor USVL246-FR2 (C. amarus) and a susceptible cultivar 'Sugar Baby' (Citrullus lanatus) to validate the utility of the markers for introgression of resistance from the wild crop relative into cultivated watermelon. Only 16 KASP markers segregated in the interspecific C. amarus/lanatus validation population. Four markers showed significant differences in the separation of genotypes based on family-mean disease severity, but together explained only 16% of the phenotypic variance. Genotypes that inherited homozygous resistant parental alleles at three KASP markers had 42% lower family-mean disease severity than homozygous susceptible genotypes. Thus, haplotype analysis was more effective at predicting the mean disease severity of families than single markers. The haplotype identified in this study will be valuable for developing Fon race 2 resistant watermelon cultivars.

Genome-Wide Association Mapping and Genomic Prediction of Fusarium Wilt Race 2 Resistance in the USDA Citrullus amarus Collection.

Fusarium wilt caused by Fusarium oxysporum f. sp. niveum (Fon) race 2 is a serious disease in watermelon and can reduce yields by 80%. Genome-wide association studies (GWAS) are a valuable tool in dissecting the genetic basis of traits. Citrullus amarus accessions (n = 120) from the USDA germplasm collection were genotyped with whole-genome resequencing, resulting in 2,126,759 single nucleotide polymorphic (SNP) markers that were utilized for GWAS. Three models were used for GWAS with the R package GAPIT. Mixed linear model (MLM) analysis did not identify any significant marker associations. FarmCPU identified four quantitative trait nucleotides (QTN) on three different chromosomes (i.e., chromosomes 1, 5, and 9), and Bayesian-information and linkage-disequilibrium iteratively nested keyway (BLINK) identified one QTN on chromosome 10 as significantly associated with Fon race 2 resistance. FarmCPU identified four QTN that explained 60% of Fon race 2 resistance, and the single QTN from BLINK explained 27%. Relevant candidate genes were found within the linkage disequilibrium (LD) blocks of these significant SNPs, including genes encoding aquaporins, expansins, 2S albumins, and glutathione S-transferases which have been shown to be involved in imparting resistance to Fusarium spp. Genomic predictions (GP) for Fon race 2 resistance using all 2,126,759 SNPs resulted in a mean prediction accuracy of 0.08 with five-fold cross-validation employing genomic best linear unbiased prediction (gBLUP) or ridge-regression best linear unbiased prediction (rrBLUP). Mean prediction accuracy with gBLUP leave-one-out cross-validation was 0.48. Thus, along with identifying genomic regions associated with Fon race 2 resistance among the accessions, this study observed prediction accuracies that were strongly influenced by population size.

Genome-Wide Association Study of Resistance to Pseudomonas syringae in the USDA Collection of Citrullus amarus.

Pseudomonas leaf spot (PLS), caused by Pseudomonas syringae pv. syringae, is an emerging disease of watermelon in the United States with the potential to severely reduce yield under humid conditions. The genetic basis of resistance to this disease is not known and no resistant germplasm is available. Because Citrullus amarus is an important reservoir of resistance genes for the cultivated watermelon, C. lanatus, we screened the United States Department of Agriculture plant introduction collection of C. amarus for resistance to PLS. Accessions (n = 117) were phenotyped for their level of resistance to PLS in two separate tests. Accession means of percent leaf area affected ranged from 1.5 to 99.4%. The broad-sense heritability for the trait was 0.51. Whole-genome resequencing generated 2,126,759 single-nucleotide polymorphisms (SNPs) which were used to perform a genome-wide association study (GWAS) aimed at discovering molecular markers for resistance. Three different models-BLINK, FarmCPU, and MLM-were included in the GWAS analyses. BLINK and FarmCPU, which are multilocus models, found eight SNPs, located on chromosomes Ca01, Ca05, Ca06, Ca08, and Ca10, that were significantly associated with resistance to PLS. Two of these SNPs were found by both BLINK and FarmCPU. The MLM model did not detect any significant associations. BLINK and FarmCPU estimated an explained phenotypic variance of 43.6 and 28.5%, respectively, for SNP S6_19327000 and 25.0 and 26.0%, respectively, for SNP S1_33362258, the two most significant SNPs found. In total, 43 candidate genes with known involvement in disease resistance were discovered within the genomic intervals of seven of the eight peak SNPs. Eleven of the candidate genes that were found have been reported to be involved in resistance to P. syringae in other plant species. Two significant SNPs were within resistance genes previously documented to play important roles of plant resistance specific to P. syringae in other pathosystems. The SNPs identified in this study will be instrumental in finding causal genes involved in PLS resistance in watermelon and developing resistant germplasm through breeding.

QTL mapping of resistance to Pseudoperonospora cubensis clade 2, mating type A1, in Cucumis melo and dual-clade marker development.

KEY MESSAGE: This is the first identification of QTLs underlying resistance in Cucumis melo to an isolate of Pseudoperonospora cubensis identified as Clade 2/mating type A1. Pseudoperonospora cubensis, causal organism of cucurbit downy mildew (CDM), causes severe necrosis and defoliation on Cucumis melo (melon). A recombinant inbred line population (N = 169) was screened against an isolate of P. cubensis (Clade 2/mating type A1) in replicated greenhouse and growth chamber experiments. SNPs (n = 5633 bins) identified in the RIL population were used for quantitative trait loci (QTL) mapping. A single major QTL on chromosome 10 (qPcub-10.3-10.4) was consistently associated with resistance across all experiments, while a second major QTL on chromosome 8 (qPcub-8.3) was identified only in greenhouse experiments. These two major QTLs were identified on the same chromosomes (8 and 10) but in different locations as two major QTLs (qPcub-8.2 and qPcub-10.1) previously identified for resistance to P. cubensis Clade 1/mating type A2. Kompetitive allele-specific PCR (KASP) markers were developed for these four major QTLs and validated in the RIL population through QTL mapping. These markers will provide melon breeders a high-throughput genotyping toolkit for development of melon cultivars with broad tolerance to CDM.

Distinct Genomic Loci Underlie Quantitative Resistance to Meloidogyne enterolobii Galling and Reproduction in Citrullus amarus.

Meloidogyne enterolobii is a virulent species of root-knot nematode that threatens watermelon (Citrullus lanatus) production in the southeastern United States. There are no known sources of root-knot nematode resistance in cultivated C. lanatus. Specific genotypes of a wild watermelon relative, C. amarus, are resistant against M. incognita but the genetics that underly this resistance are still unknown and it is not clear that this same resistance will be effective against M. enterolobii. To identify and characterize new sources of resistance to M. enterolobii, we screened 108 diverse C. amarus lines alongside a susceptible C. lanatus cultivar (Charleston Gray) for resistance against M. enterolobii. Different C. amarus genotypes ranged from resistant to susceptible for the three resistance phenotypes measured; mean percent root system galled ranged from 10 to 73%, mean egg mass counts ranged from 0.3 to 64.5, and mean eggs per gram of root ranged from 326 to 146,160. We used each of these three resistance phenotypes combined with whole-genome resequencing data to conduct a genome-wide association scan that identified significant associations between M. enterolobii resistance and 11 single-nucleotide polymorphisms (SNPs) within the C. amarus genome. Interestingly, SNPs associated with reduced galling and egg masses were located within a single quantitative trait locus (QTL) on chromosome Ca03, while reductions in nematode eggs per gram of root were associated with separate QTL on chromosomes Ca04 and Ca08. The results of this study suggest that multiple genes are involved with M. enterolobii resistance in C. amarus and the SNPs identified will assist with efforts to breed for M. enterolobii resistance in watermelon.

Graph-based pangenomics maximizes genotyping density and reveals structural impacts on fungal resistance in melon.

The genomic sequences segregating in experimental populations are often highly divergent from the community reference and from one another. Such divergence is problematic under various short-read-based genotyping strategies. In addition, large structural differences are often invisible despite being strong candidates for causal variation. These issues are exacerbated in specialty crop breeding programs with fewer, lower-quality sequence resources. Here, we examine the benefits of complete genomic information, based on long-read assemblies, in a biparental mapping experiment segregating at numerous disease resistance loci in the non-model crop, melon (Cucumis melo). We find that a graph-based approach, which uses both parental genomes, results in 19% more variants callable across the population and raw allele calls with a 2 to 3-fold error-rate reduction, even relative to single reference approaches using a parent genome. We show that structural variation has played a substantial role in shaping two Fusarium wilt resistance loci with known causal genes. We also report on the genetics of powdery mildew resistance, where copy number variation and local recombination suppression are directly interpretable via parental genome alignments. Benefits observed, even in this low-resolution biparental experiment, will inevitably be amplified in more complex populations.

Genome-Wide Association Analysis of Resistance to Pseudoperonospora cubensis in Citron Watermelon.

Cultivated sweet watermelon (Citrullus lanatus) is an important vegetable crop for millions of people around the world. There are limited sources of resistance to economically important diseases within C. lanatus, whereas C. amarus has a reservoir of traits that can be exploited to improve C. lanatus for resistance to biotic and abiotic stresses. Cucurbit downy mildew (CDM), caused by Pseudoperonospora cubensis, is an emerging threat to watermelon production. We screened 122 C. amarus accessions for resistance to CDM over two tests (environments). The accessions were genotyped by whole-genome resequencing to generate 2,126,759 single nucleotide polymorphic (SNP) markers. A genome-wide association study was deployed to uncover marker-trait associations and identify candidate genes underlying resistance to CDM. Our results indicate the presence of wide phenotypic variability (1.1 to 57.8%) for leaf area infection, representing a 50.7-fold variation for CDM resistance across the C. amarus germplasm collection. Broad-sense heritability estimate was 0.55, implying the presence of moderate genetic effects for resistance to CDM. The peak SNP markers associated with resistance to P. cubensis were located on chromosomes Ca03, Ca05, Ca07, and Ca11. The significant SNP markers accounted for up to 30% of the phenotypic variation and were associated with promising candidate genes encoding leucine-rich repeat receptor-like protein kinase and the WRKY transcription factor. This information will be useful in understanding the genetic architecture of the P. cubensis-Citrullus spp. patho-system as well as development of resources for genomics-assisted breeding for resistance to CDM in watermelon.

Quantitative Trait Loci Mapping of Resistance to Powdery Mildew Race 1 in a Recombinant Inbred Line Population of Melon.

Powdery mildew, caused by the fungus Podosphaera xanthii, is one of the most important diseases of melon. Although there are several pathogenic races of P. xanthii, race 1 is the predominant race in South Carolina and in other parts of the United States. We used a densely genotyped recombinant inbred line melon population for traditional quantitative trait loci (QTL) mapping, to identify two major (qPx1-5 and qPx1-12) and two minor (qPx1-4 and qPx1-10) QTLs (named according to race - chromosome number) associated with resistance to P. xanthii race 1. QTL mapping of disease severity in multiple tissues (hypocotyl, cotyledons, true leaves, and stems) identified the same genetic basis of resistance in all tissue types. Whole-genome resequencing of the parents was used for marker development across the major QTLs and functional annotation of single nucleotide polymorphisms (SNPs) for candidate gene analysis. Kompetitive allele-specific PCR (KASP) markers were tightly linked to the QTL peaks of qPx1-5 (pm1-5_25329892, pm1-5_25461503 and pm1-5_25625375) and qPx1-12 (pm1-12_22848920 and pm1-12_22904659) in the population and will enable efficient marker-assisted introgression of powdery mildew resistance into improved germplasm. Candidate genes were identified in both major QTL intervals that encode putative R genes with missense mutations between the parents. The candidate genes provide targets for future breeding efforts and a fundamental examination of resistance to powdery mildew in melon.

QTL mapping of resistance to Pseudoperonospora cubensis clade 1, mating type A2, in Cucumis melo.

KEY MESSAGE: This is the first identification of QTLs underlying resistance to Pseudoperonospora cubensis in Cucumis melo using a genetically characterized isolate. Pseudoperonospora cubensis, causal organism of cucurbit downy mildew (CDM), is one of the largest threats to cucurbit production in the eastern USA. Currently, no Cucumis melo (melon) cultivars have significant levels of resistance. Additionally, little is understood about the genetic basis of resistance in C. melo. Recombinant inbred lines (RILs; N = 169) generated from a cross between the resistant melon breeding line MR-1 and susceptible cultivar Ananas Yok'neam were phenotyped for CDM resistance in both greenhouse and growth chamber studies. A high-density genetic linkage map with 5,663 binned SNPs created from the RIL population was utilized for QTL mapping. Nine QTLs, including two major QTLs, were associated with CDM resistance. Of the major QTLs, qPcub-10.1 was stable across growth chamber and greenhouse tests, whereas qPcub-8.2 was detected only in growth chamber tests. qPcub-10.1 co-located with an MLO-like protein coding gene, which has been shown to confer resistance to powdery mildew and Phytophthora in other plants. This is the first screening of C. melo germplasm with a genetically characterized P. cubensis isolate.

QTL Mapping and Marker Development for Tolerance to Sulfur Phytotoxicity in Melon (Cucumis melo).

Elemental sulfur is an effective, inexpensive fungicide for many foliar pathogens, but severe phytotoxicity prohibits its use on many melon varieties. Sulfur phytotoxicity causes chlorosis and necrosis of leaf tissue, leading to plant death in the most sensitive lines, while other varieties have little to no damage. A high-density, genotyping-by-sequencing (GBS)-based genetic map of a recombinant inbred line (RIL) population segregating for sulfur tolerance was used for a quantitative trait loci (QTL) mapping study of sulfur phytotoxicity in melon. One major (qSulf-1) and two minor (qSulf-8 and qSulf-12) QTL were associated with sulfur tolerance in the population. The development of Kompetitive Allele-Specific PCR (KASP) markers developed across qSulf-1 decreased the QTL interval from 239 kb (cotyledons) and 157 kb (leaves) to 97 kb (both tissues). The markers were validated for linkage to sulfur tolerance in a set of melon cultivars. These KASP markers can be incorporated into melon breeding programs for introgression of sulfur tolerance into elite melon germplasm.

QTL mapping of resistance to Fusarium oxysporum f. sp. niveum race 2 and Papaya ringspot virus in Citrullus amarus.

KEY MESSAGE: A Citrullus amarus mapping population segregating for resistance to Fusarium oxysporum f. sp. niveum race 2 and Papaya ringspot virus was used to identify novel QTL, important for the improvement in watermelon disease resistance. Multiple disease screens of the USDA Citrullus spp. germplasm collection have highlighted the value of Citrullus amarus (citron melon or wild watermelon) as a resource for enhancing modern watermelon cultivars (Citrullus lanatus) with resistance to a broad range of fungal, bacterial and viral diseases of watermelon. We have generated a genetic population of C. amarus segregating for resistance to two important watermelon diseases: Fusarium wilt (caused by the fungus Fusarium oxysporum f. sp. niveum; Fon race 2) and Papaya ringspot virus-watermelon strain (PRSV-W). QTL mapping of Fon race 2 resistance identified seven significant QTLs, with the major QTL representing a novel genetic source of resistance and an opportunity for gene pyramiding. A single QTL was associated with resistance to PRSV-W, which adhered to expectations of a prior study indicating a single-gene recessive inheritance in watermelon. The resistance loci identified here provide valuable genetic resources for introgression into cultivated watermelon for the improvement in disease resistance.

QTL Mapping Identifies Novel Source of Resistance to Fusarium Wilt Race 1 in Citrullus amarus.

Fusarium wilt race 1, caused by the soilborne fungus Fusarium oxysporum Schlechtend.: Fr. f. sp. niveum (E.F. Sm.) W.C. Snyder & H.N. Hans (Fon), is a major disease of watermelon (Citrullus lanatus) in the United States and throughout the world. Although Fusarium wilt race 1 resistance has been incorporated into several watermelon cultivars, identification of additional genetic sources of resistance is crucial if a durable and sustainable level of resistance is to be continued over the years. We conducted a genetic mapping study to identify quantitative trait loci (QTLs) associated with resistance to Fon race 1 in segregating populations (F2:3 and recombinant inbred lines) of Citrullus amarus (citron melon) derived from the Fon race 1 resistant and susceptible parents USVL246-FR2 and USVL114, respectively. A major QTL (qFon1-9) associated with resistance to Fon race 1 was identified on chromosome 9 of USVL246-FR2. This discovery provides a novel genetic source of resistance to Fusarium wilt race 1 in watermelon and, thus, an additional host-resistance option for watermelon breeders to further the effort to mitigate this serious phytopathogen.

Genome-Wide Association Study of Resistance to Xanthomonas gardneri in the USDA Pepper (Capsicum) Collection.

Bacterial spot Xanthomonas species cause significant disease outbreaks on tomato and pepper in tropical and subtropical regions throughout the world. Host resistance has been one of the key components of integrated disease management approaches to mitigate plant pathogens. Although a number of resistance genes have been identified in pepper against bacterial spot xanthomonads, emergence of bacterial spot pathogen variants capable of overcoming these sources and changing pathogen distributions reinforce the importance of identifying novel candidates to incorporate into breeding programs. We conducted a genome-wide association study (GWAS) on a diverse U.S. Department of Agriculture collection of pepper germplasm including different species of Capsicum to identify novel sources of disease resistance against a highly virulent X. gardneri strain isolated from a recent outbreak. GWAS identified highly significant single nucleotide polymorphisms associated with defoliation in response to infection with X. gardneri. Functionally relevant candidate genes encoded products involved in disease resistance/susceptibility, hormone signaling, and basal resistance against multiple pathogens in various host-pathogen systems. The X. gardneri-resistant genotypes and quantitative trait loci identified in this study provide alleles that could be used for a resistance gene pyramiding effort against different species of bacterial spot xanthomonads in pepper.

QTL mapping of resistance to bacterial fruit blotch in Citrullus amarus.

KEY MESSAGE: Six QTLs were associated with affected leaf area in response to inoculation with Acidovorax citrulli in a recombinant inbred line population of Citrullus amarus. Acidovorax citrulli, the causal agent of bacterial fruit blotch (BFB) of cucurbits, has the potential to devastate production of watermelon and other cucurbits. Despite decades of research on host-plant resistance to A. citrulli, no germplasm has been found with immunity and only a few sources with various levels of BFB resistance have been identified, but the genetic basis of resistance in these watermelon sources are not known. Most sources of resistance are plant introductions of Citrullus amarus (citron melon), a closely related species that crosses readily with cultivated watermelon (Citrullus lanatus L.). In this study, we evaluated a recombinant inbred line population (N = 200), derived from a cross between BFB-resistant (USVL246-FR2) and BFB-susceptible (USVL114) C. amarus lines, for foliar resistance to A. citrulli in three replicated greenhouse trials. We found the genetics of BFB resistance to be complicated by strong environmental influence, low heritability and significant genotype-by-environment interactions. QTL mapping of affected leaf area identified six QTL that each explained between 5 and 15% of the variation in BFB resistance in the population. This study represents the first identification of QTL associated with resistance to A. citrulli in any cucurbit.

Genetic Diversity and Population Structure of the USDA Sweetpotato (Ipomoea batatas) Germplasm Collections Using GBSpoly.

Sweetpotato (Ipomoea batatas) plays a critical role in food security and is the most important root crop worldwide following potatoes and cassava. In the United States (US), it is valued at over $700 million USD. There are two sweetpotato germplasm collections (Plant Genetic Resources Conservation Unit and US Vegetable Laboratory) maintained by the USDA, ARS for sweetpotato crop improvement. To date, no genome-wide assessment of genetic diversity within these collections has been reported in the published literature. In our study, population structure and genetic diversity of 417 USDA sweetpotato accessions originating from 8 broad geographical regions (Africa, Australia, Caribbean, Central America, Far East, North America, Pacific Islands, and South America) were determined using single nucleotide polymorphisms (SNPs) identified with a genotyping-by-sequencing (GBS) protocol, GBSpoly, optimized for highly heterozygous and polyploid species. Population structure using Bayesian clustering analyses (STRUCTURE) with 32,784 segregating SNPs grouped the accessions into four genetic groups and indicated a high degree of mixed ancestry. A neighbor-joining cladogram and principal components analysis based on a pairwise genetic distance matrix of the accessions supported the population structure analysis. Pairwise FST values between broad geographical regions based on the origin of accessions ranged from 0.017 (Far East - Pacific Islands) to 0.110 (Australia - South America) and supported the clustering of accessions based on genetic distance. The markers developed for use with this collection of accessions provide an important genomic resource for the sweetpotato community, and contribute to our understanding of the genetic diversity present within the US sweetpotato collection and the species.

Construction of a genome-anchored, high-density genetic map for melon (Cucumis melo L.) and identification of Fusarium oxysporum f. sp. melonis race 1 resistance QTL.

KEY MESSAGE: Four QTLs and an epistatic interaction were associated with disease severity in response to inoculation with Fusarium oxysporum f. sp. melonis race 1 in a recombinant inbred line population of melon. The USDA Cucumis melo inbred line, MR-1, harbors a wealth of alleles associated with resistance to several major diseases of melon, including powdery mildew, downy mildew, Alternaria leaf blight, and Fusarium wilt. MR-1 was crossed to an Israeli cultivar, Ananas Yok'neam, which is susceptible to all of these diseases, to generate a recombinant inbred line (RIL) population of 172 lines. In this study, the RIL population was genotyped to construct an ultra-dense genetic linkage map with 5663 binned SNPs anchored to the C. melo genome and exhibits the overall high quality of the assembly. The utility of the densely genotyped population was demonstrated through QTL mapping of a well-studied trait, resistance to Fusarium wilt caused by Fusarium oxysporum f. sp. melonis (Fom) race 1. A major QTL co-located with the previously validated resistance gene Fom-2. In addition, three minor QTLs and an epistatic interaction contributing to Fom race 1 resistance were identified. The MR-1 × AY RIL population provides a valuable resource for future QTL mapping studies and marker-assisted selection of disease resistance in melon.

Quantitative trait loci mapping of heat tolerance in broccoli (Brassica oleracea var. italica) using genotyping-by-sequencing.

KEY MESSAGE: Five quantitative trait loci and one epistatic interaction were associated with heat tolerance in a doubled haploid population of broccoli evaluated in three summer field trials. Predicted rising global temperatures due to climate change have generated a demand for crops that are resistant to yield and quality losses from heat stress. Broccoli (Brassica oleracea var. italica) is a cool weather crop with high temperatures during production decreasing both head quality and yield. Breeding for heat tolerance in broccoli has potential to both expand viable production areas and extend the growing season but breeding efficiency is constrained by limited genetic information. A doubled haploid (DH) broccoli population segregating for heat tolerance was evaluated for head quality in three summer fields in Charleston, SC, USA. Multiple quantitative trait loci (QTL) mapping of 1,423 single nucleotide polymorphisms developed through genotyping-by-sequencing identified five QTL and one positive epistatic interaction that explained 62.1% of variation in heat tolerance. The QTL identified here can be used to develop markers for marker-assisted selection and to increase our understanding of the molecular mechanisms underlying plant response to heat stress.

A GBS-SNP-based linkage map and quantitative trait loci (QTL) associated with resistance to Fusarium oxysporum f. sp. niveum race 2 identified in Citrullus lanatus var. citroides.

KEY MESSAGE: A major QTL for resistance to Fusarium oxysporum f. sp. niveum race 2 was mapped to a narrow 1.2 Mb interval using a high-density GBS-SNP linkage map, the first map of Citrullus lanatus var. citroides. Fusarium wilt, a fungal disease caused by Fusarium oxysporum f. sp. niveum (Fon), devastates watermelon crop production worldwide. Several races, which are differentiated by host range, of the pathogen exist. Resistance to Fon race 2, a particularly virulent strain prevalent in the United States, does not exist in edible cultivars of the sweet cultivated watermelon Citrullus lanatus var. lanatus (Cll) and has been well described in a few plant introductions of the wild subspecies of watermelon, C. lanatus var. citroides (Clc). Clc provides a vital source of genetic diversity, as well as resistance to numerous diseases. Unfortunately, both genetic diversity and disease resistance are lacking in Cll due to the narrow genetic base. Despite the importance of Clc to continued watermelon improvement, intra-variety genetic studies are lacking. Here, we present the first Clc genetic linkage map, generated with 2495 single nucleotide polymorphisms developed through genotyping-by-sequencing, and use it to identify quantitative trait loci associated with Fon race 2 resistance. Multiple QTL mapping in a Clc F2:3 population (N = 173) identified one major and four minor QTL. The major QTL explained 43% of the variation in Fon race 2 resistance and was delimited to a 1.2-Mb interval on chromosome 9, a region spanning 44 genes.

Genome-Wide Association Study in Arabidopsis thaliana of Natural Variation in Seed Oil Melting Point: A Widespread Adaptive Trait in Plants.

Seed oil melting point is an adaptive, quantitative trait determined by the relative proportions of the fatty acids that compose the oil. Micro- and macro-evolutionary evidence suggests selection has changed the melting point of seed oils to covary with germination temperatures because of a trade-off between total energy stores and the rate of energy acquisition during germination under competition. The seed oil compositions of 391 natural accessions of Arabidopsis thaliana, grown under common-garden conditions, were used to assess whether seed oil melting point within a species varied with germination temperature. In support of the adaptive explanation, long-term monthly spring and fall field temperatures of the accession collection sites significantly predicted their seed oil melting points. In addition, a genome-wide association study (GWAS) was performed to determine which genes were most likely responsible for the natural variation in seed oil melting point. The GWAS found a single highly significant association within the coding region of FAD2, which encodes a fatty acid desaturase central to the oil biosynthesis pathway. In a separate analysis of 15 a priori oil synthesis candidate genes, 2 (FAD2 and FATB) were located near significant SNPs associated with seed oil melting point. These results comport with others' molecular work showing that lines with alterations in these genes affect seed oil melting point as expected. Our results suggest natural selection has acted on a small number of loci to alter a quantitative trait in response to local environmental conditions.

Genome-Wide Association Study of Arabidopsis thaliana Identifies Determinants of Natural Variation in Seed Oil Composition.

The renewable source of highly reduced carbon provided by plant triacylglycerols (TAGs) fills an ever increasing demand for food, biodiesel, and industrial chemicals. Each of these uses requires different compositions of fatty acid proportions in seed oils. Identifying the genes responsible for variation in seed oil composition in nature provides targets for bioengineering fatty acid proportions optimized for various industrial and nutrition goals. Here, we characterized the seed oil composition of 391 world-wide, wild accessions of Arabidopsis thaliana, and performed a genome-wide association study (GWAS) of the 9 major fatty acids in the seed oil and 4 composite measures of the fatty acids. Four to 19 regions of interest were associated with the seed oil composition traits. Thirty-four of the genes in these regions are involved in lipid metabolism or transport, with 14 specific to fatty acid synthesis or breakdown. Eight of the genes encode transcription factors. We have identified genes significantly associated with variation in fatty acid proportions that can be used as a resource across the Brassicaceae. Two-thirds of the regions identified contain candidate genes that have never been implicated in lipid metabolism and represent potential new targets for bioengineering.