Optimization of AFLP for extremely large genomes over 70 Gb

Here, we present an improved amplified fragment length polymorphism (AFLP) protocol using restriction enzymes (AscI and SbfI) that recognize 8‐base pair sequences to provide alternative optimization suitable for species with a genome size over 70 Gb. This cost‐effective optimization massively reduces the number of amplified fragments using only +3 selective bases per primer during selective amplification. We demonstrate the effects of the number of fragments and genome size on the appearance of nonidentical comigrating fragments (size homoplasy), which has a negative impact on the informative value of AFLP genotypes. We also present various reaction conditions and their effects on reproducibility and the band intensity of the extremely large genome of Viscum album. The reproducibility of this octo‐cutter protocol was calculated using several species with genome sizes ranging from 1 Gb (Carex panicea) to 76 Gb (V. album). The improved protocol also succeeded in detecting high intraspecific variability in species with large genomes (V. album, Galanthus nivalis and Pinus pumila).     

Flavonoid evidence for allopolyploidy in the Chenopodium album aggregate (Amaranthaceae)

The flavonoid chemistry of 16 species of Chenopodium was investigated, with an emphasis on C. album and its relatives. The chief compounds seen were 3-O-glycosides of quercetin, kaempferol and isorhamnetin. The latter two classes show a mutually exclusive distribution among the diploid and all but one tetraploid species. C. polyspermum is unusual in possessing O-methylation at the 4-, 6- and 7-positions, and C. murale is the only taxon to have 7-O-glycosylation. Acylated glycosides are common. C. album and related hexaploid taxa display a single flavonoid profile, providing no support for the recognition of more than one species. The hexaploid flavonoid profile represents an almost perfect summation of those of the diploids, C. suecicum and C. ficifolium. This apparent additive inheritance supports the hypothesis that these species (or taxa very similar to them) are involved in the ancestry of C. album. Chromosome numbers are reported for 14 of the species.     

<Emphasis Type="Italic">Chenopodium</Emphasis>: a prospective plant for phytoextraction

Leaf samples were collected from 40 accessions of Chenopodium spp. and assessed for six heavy metals (Fe, Zn, Cu, Ni, Cr and Cd) accumulation to explore the use of Chenopodium for phytoextraction of heavy metals. The results suggest that Chenopodium spp. have the ability to accumulate large quantities of heavy metals in the leaf tissues even when they are present in low concentrations in the soil. C. quinoa is a better accumulator of Ni, Cr and Cd than the rest of the species, while C. album accessions are good copper accumulators. Bioconcentration factor for chromium ranged from 0.36 (C. album “Chandanbathua”) to 6.57 (C. quinoa Ames 13719) with 13 accessions of C. quinoa scoring above the mean value. High heritability coupled with high genetic advance was recorded for Ni, Cr and Cd, which indicated a major role of additive gene action in the inheritance of these characters. Zinc showed significant positive association with iron (0.351**), nickel (0.659**), chromium (0.743**) and cadmium (0.288**). Nickel was significantly and negatively associated with copper (−0.663**), while it was positively and significantly correlated with chromium (0.682**) and cadmium (0.461**). Considering the accumulation efficiency of Chenopodium spp. with respect to heavy metals, this genus should be further explored for decontamination of metal polluted soils, with plant breeding playing an important role in evolving new plant types with higher capacity of heavy metal accumulation.     

Transition from C3 to proto-Kranz to C3–C4 intermediate type in the genus Chenopodium (Chenopodiaceae)

The Chenopodiaceae is one of the families including C₄ species among eudicots. In this family, the genus Chenopodium is considered to include only C₃ species. However, we report here a transition from C₃ photosynthesis to proto-Kranz to C₃–C₄ intermediate type in Chenopodium. We investigated leaf anatomical and photosynthetic traits of 15 species, of which 8 species showed non-Kranz anatomy and a CO₂ compensation point (Γ) typical of C₃ plants. However, 5 species showed proto-Kranz anatomy and a C₃-like Γ, whereas C. strictum showed leaf anatomy and a Γ typical of C₃–C₄ intermediates. Chenopodium album accessions examined included both proto-Kranz and C₃–C₄ intermediate types, depending on locality. Glycine decarboxylase, a key photorespiratory enzyme that is involved in the decarboxylation of glycine, was located predominantly in the mesophyll (M) cells of C₃ species, in both M and bundle-sheath (BS) cells in proto-Kranz species, and exclusively in BS cells in C₃–C₄ intermediate species. The M/BS tissue area ratio, number of chloroplasts and mitochondria per BS cell, distribution of these organelles to the centripetal region of BS cells, the degree of inner positioning (vacuolar side of chloroplasts) of mitochondria in M cells, and the size of BS mitochondria also changed with the change in glycine decarboxylase localization. All Chenopodium species examined were C₃-like regarding activities and amounts of C₃ and C₄ photosynthetic enzymes and δ¹³C values, suggesting that these species perform photosynthesis without contribution of the C₄ cycle. This study demonstrates that Chenopodium is not a C₃ genus and is valuable for studying evolution of C₃–C₄ intermediates.

     

Measures of ecological association

Summary The relationships of photosynthetic characteristics to the competitive interactions of a C₃ plant, Chenopodium album, and a C₄ plant, Amaranthis retroflexus, were investigated in different temperature and water supply regimes. Both species had similar photosynthetic rates at 25°C, but at higher temperatures, Amaranthus had substantially greater rates than Chenopodium. Conversely, at lower temperatures, Chenopodium had an advantage. The competitive abilities in mixtures exhibited a close parallel to the photosynthetic performances with Amaranthus having an advantage at high temperatures and Chenopodium an advantage at low temperatures. These competitive outcomes were determined primarily by differences in relative growth rates prior to canopy closure. In the respective, temperature regimes, the species having the highest photosynthetic rate, which resulted an more rapid growth, overtopped and shaded the other species at the time of canopy closure. These results demonstrate that differences in photosynthetic temperature response between C₄ and C₃ plants can be an important determinant in competitive interactions, but at least in this case, the influence is primarily through, events prior to the actual initiation of competition.In contrast to temperature, growth of the plants under limited water supply had no influence on the competitive interactions. Thus, the presence of the C₄ pathway alone was not sufficient to yield a competitive advantage over the C₃ species under water limited conditions.     

Genome size variation in Orchidaceae subfamily Apostasioideae: filling the phylogenetic gap

With more than 160‐fold variation, Orchidaceae are currently the most diverse angiosperm family with respect to the amount of nuclear DNA. This study provides first genome size estimates for approximately 50% of species currently recognized in subfamily Apostasioideae, which is sister to the other four orchid subfamilies. The estimated 1C‐values range from 0.38 pg in Apostasia nuda to 5.96 pg in Neuwiedia zollingeri var. javanica, a nearly 16‐fold range. The two genera show non‐overlapping genome sizes, with those in Apostasia being distinctly smaller than those in Neuwiedia. In fact, most Apostasia spp. are at the lower end of the range of orchid C‐values. Observed discontinuities in DNA amounts in genera most probably reflect interspecific variation in ploidy. In addition to ploidy heterogeneity in N. zollingeri var. javanica, intraspecific variation in genome size (up to 17.7%) was also detected in some species; this can be plausibly related to the incidence of different geographical variants or unrecognized taxonomic heterogeneity. The AT content varied from 62.6 to 66.0%, which is in the upper range recorded for angiosperms. The genome size data obtained in this study fill a major phylogenetic gap in Orchidaceae and show that (very) small genomes prevail in subfamily Apostasioideae. © 2013 The Linnean Society of London     

Morphological and molecular analyses support the existence of host-specific Peronospora species infecting Chenopodium

About 20 species of Peronospora have been reported to cause downy mildew on Chenopodium, but, particularly in plant pathology literature, only one species, P. farinosa, is considered to be involved. We performed sequence analysis of the ITS rDNA to reveal the phylogenetic relationships of Peronospora specimens from five species of Chenopodium, viz. C. album, C. ambrosioides, C. bonus-henricus, C. hybridum, and C. polyspermum. The five clades corresponded to particular Chenopodium species, and showed a high level of sequence divergence. Differences in the morphology of the conidia and ultimate branchlets also supported the separation of the five groups at the host species level. These results suggest that the names P. variabilis, P. boni-henrici, P. chenopodii, and P. chenopodii-polyspermi should be used for the four downy mildew pathogens specific to C. album, C. bonus-henricus, C. hybridum, and C. polyspermum, respectively. The Peronospora on C. ambrosioides was found to be an independent species.     

Systematics and biogeography of the ant genus Crematogaster Lund subgenus Orthocrema Santschi in Asia (Hymenoptera: Formicidae)

The subgenus Orthocrema of the ant genus Crematogaster is a well defined group and diverse in the tropical Asia. Its systematics has remained poorly understood because of a lack of modern revisionary work. Crematogaster (Orthocrema) is revised for the Asian region, and 27 species including ten new species are recognized. Five species groups: the C. baduvi group (4 spp.); the C. binghamii group (3 spp.); the C. biroi group (10 spp.); the C. moatensis group (1 sp.); the C. quadriruga group (9 spp.) are established based on worker caste morphology. A key to Asian species of the subgenus Orthocrema based on the worker caste is given. Phylogenetic relationships of Asian Orthocrema are analyzed. The analysis revealed that the C. baduvi‐, C. binghamii‐, and C. biroi groups are monophyletic, and that the allopatric distribution patterns of closely related species imply that Asian Orthocrema is composed of relatively young taxa. There have been at least three west‐to‐east dispersal events across Wallace's line in the C. baduvi‐, C. quadriruga‐ and C. biroi groups. © 2016 The Linnean Society of London     

Floral specialization for different pollinators and divergent use of the same pollinator among co‐occurring Impatiens species (Balsaminaceae) from Southeast Asia

Floral variation among closely related species is thought to often reflect differences in pollination systems. Flowers of the large genus Impatiens are characterized by extensive variation in colour, shape and size and in anther and stigma positioning, but studies of their pollination ecology are scarce and most lack a comparative context. Consequently, the function of floral diversity in Impatiens remains enigmatic. This study documents floral variation and pollination of seven co‐occurring Impatiens spp. in the Southeast Asian diversity hotspot. To assess whether floral trait variation reflects specialization for different pollination systems, we tested whether species depend on pollinators for reproduction, identified animals that visit flowers, determined whether these visitors play a role in pollination and quantified and compared key floral traits, including floral dimensions and nectar characteristics. Experimental exclusion of insects decreased fruit and seed set significantly for all species except I. muscicola, which also received almost no visits from animals. Most species received visits from several animals, including bees, birds, butterflies and hawkmoths, only a subset of which were effective pollinators. Impatiens psittacina, I. kerriae, I. racemosa and I. daraneenae were pollinated by bees, primarily Bombus haemorrhoidalis. Impatiens chiangdaoensis and I. santisukii had bimodal pollination systems which combined bee and lepidopteran pollination. Floral traits differed significantly among species with different pollination systems. Autogamous flowers were small and spurless, and did not produce nectar; bee‐pollinated flowers had short spurs and large floral chambers with a wide entrance; and bimodally bee‐ and lepidopteran‐pollinated species had long spurs and a small floral chamber with a narrow entrance. Nectar‐producing species with different pollination systems did not differ in nectar volume and sugar concentration. Despite the high frequency of bee pollination in co‐occurring species, individuals with a morphology suggestive of hybrid origin were rare. Variation in floral architecture, including various forms of corolla asymmetry, facilitates distinct, species‐specific pollen‐placement on visiting bees. Our results show that floral morphological diversity among Impatiens spp. is associated with both differences in functional pollinator groups and divergent use of the same pollinator. Non‐homologous mechanisms of floral asymmetry are consistent with repeated independent evolution, suggesting that competitive interactions among species with the same pollination system have been an important driver of floral variation among Impatiens spp.     

Phytochemical and Cytogenetic Characterization of Centaurea solstitialis L. (Asteraceae) from Croatia

The cytogenetic characterization of Centaurea solstitialis L. (Asteraceae) showed a chromosome number of 2n = 16. Karyotype is composed by four pairs of metacentric, two pairs of submetacentric and two pairs of subtelocentric chromosomes. Physical mapping of two rDNA probes revealed two loci of 35S and one locus of 5S rRNA genes. Chromomycin fluorochrome banding revealed that all rDNA loci were GC rich. The genome size (2C‐value) of 1.95 pg classes this species in the group of very small genomes. Chemical composition of C. solstitialis volatile oil (VO) from Croatia, studied with gas chromatography–mass spectrometry showed dominant components as it follows: hexadecanoic acid, α‐linolenic acid, germacrene D and heptacosane. Antioxidant capacity, measured by ferric reducing power assay and 2,2‐diphenyl‐1‐picrylhydrazyl methods, as well as inhibition of acetyl‐ and butyrylcholinesterase of VO was lower comparing to a standard solutions. Volatile oil tested with disc diffusion method showed good inhibitory potential against Pseudomonas aeruginosa, Escherichia coli and all tested fungi: Candida albicans, Penicillium funiculosum and Aspergillus fumigatus. The microdilution method showed best activity against Chronobacter sakazakii and A. fumigatus.     

Immunoassay‐based Techniques for Early and Specific Detection of Latent Postharvest Anthracnose in Mango

The postharvest anthracnose pathogen Colletotrichum gloeosporioides inciting latent or quiescent infection of mango was detected in early stages using immunoassay methods. Twenty‐five pathotypes isolated from different agroclimatic zones of Tamil Nadu, Karnataka and Pondicherry, India, revealed the variation in protein profile analysis (SDS‐PAGE). The polyclonal antibodies (PCA) were raised against the unfractioned mycelial protein (UMP) and a 40‐kDa polypeptide present in all pathotypes. Standardization of antigen and antiserum dilutions revealed that an antigen dilution of 1 : 200 (protein concentration of 20 μg/ml) and antiserum dilution of 1 : 100 (protein concentration of 40 μg/ml raised against UMP) and 1 : 200 (protein concentration of 20 μg/ml raised against 40 kDa polypeptide) was found to be optimum for the detection of anthracnose pathogen. Both antisera detected the C. gloeosporioides antigen in enzyme‐linked immunosorbent assays (ELISAs), dot immunobinding assays (DIBAs) and Western blots. The specificity in reaction was compared by isolating other Colletotrichum spp. from various hosts viz., C. lindemuthianum (beans), C. falcatum (sugarcane), C. musae (banana), C. capsici (chillies) and Botryodiplodia theobromae (mango). The antisera generated against UMP revealed the cross‐reaction with other host isolates and mango stem end rot pathogen (B. theobromae). The PCA raised against 40‐kDa polypeptide exhibited the specific reaction with C. gloeosporioides isolates in all the immunoassay techniques. By utilizing both PCA, the presence of latent infection was observed in healthy‐looking leaves, flowers and fruits in orchard conditions. The fruit tissues recorded high absorbance values followed by flowers and leaves in all the detection methods. The ELISA technique was also useful in assessing the pathogen inoculum at various biocontrol formulations sprayed mango trees under field conditions. The fluorescent pseudomonad strains mixture (KFP1 + FP7) amended with chitin sprayed at 30‐day intervals revealed the significant reduction in pathogen load than other formulations and unsprayed control.     

Qualitative and quantitative analysis of the genomes and chromosomes of spider monkeys (Primates: Atelidae)

Heterochromatin distribution and chromosomal rearrangements have been proposed as the main sources of karyotype differences among species of Neotropical primates. This variability suggests that there could be differences at other smaller‐scale levels of DNA organization as well. In particular, quantitative differences between genomes result from gains and losses of individual DNA segments, and may result in varying genome sizes (C‐values) among species. In this work, we studied the genomes of 23 individuals from four species in the genus Ateles (Primates: Platyrrhini): A. chamek, A. paniscus, A. belzebuth, and A. geoffroyi. We analyzed genome size and its relationship with the presence of chromosomal rearrangements and patterns of heterochromatin distribution. The C‐value presented in this work for Ateles chamek is the first estimate for this species (3.09 ± 0.23 pg), whereas our estimates for A. belzebuth (2.88 ± 0.06 pg) and A. geoffroyi (3.19 ± 0.24 pg) differed from those previously published. Fluorescent in situ hybridization (FISH) and interspecies comparativegenomic hybridization (iCGH) analyses revealed that differences in genome size among species relate to localized blocks in both heterochromatic and euchromatic regions, the latter of which appear to be genetically unstable. There were also quantitative differences in Y chromosome content. It remains to be seen whether the chromosomal characteristics of Ateles here discussed are common to platyrrhine monkeys, but it is clear that these monkeys exhibit some intriguing genomic features worthy of additional exploration.     

Effects of growth temperature and nitrogen nutrition on expression of C3–C4 intermediate traits in Chenopodium album

Proto-Kranz plants represent an initial phase in the evolution from C₃ to C₃–C₄ intermediate to C₄ plants. The ecological and adaptive aspects of C₃–C₄ plants would provide an important clue to understand the evolution of C₃–C₄ plants. We investigated whether growth temperature and nitrogen (N) nutrition influence the expression of C₃–C₄ traits in Chenopodium album (proto-Kranz) in comparison with Chenopodium quinoa (C₃). Plants were grown during 5 weeks at 20 or 30 °C under standard or low N supply levels (referred to as 20SN, 20LN, 30SN, and 30LN). Net photosynthetic rate and leaf N content were higher in 20SN and 30SN plants than in 20LN and 30LN plants of C. album but did not differ among growth conditions in C. quinoa. The CO₂ compensation point (Γ) of C. album was lowest in 30LN plants (36 µmol mol^–1), highest in 20SN plants (51 µmol mol^–1), and intermediate in 20LN and 30SN plants, whereas Γ of C. quinoa did not differ among the growth conditions (51–52 µmol mol^–1). The anatomical structure of leaves was not considerably affected by growth conditions in either species. However, ultrastructural observations in C. album showed that the number of mitochondria per mesophyll or bundle sheath (BS) cell was lower in 20LN and 30LN plants than in 20SN and 30SN plants. Immunohistochemical observations revealed that lower accumulation level of P-protein of glycine decarboxylase (GDC-P) in mesophyll mitochondria than in BS mitochondria is the major factor causing the decrease in Γ values in C. album plants grown under low N supply and high temperature. These results suggest that high growth temperature and low N supply lead to the expression of C₃–C₄ traits (the reduction of Γ) in the proto-Kranz plants of C. album through the regulation of GDC-P expression.

     

The Chimonanthus salicifolius genome provides insight into magnoliid evolution and flavonoid biosynthesis

Chimonanthus salicifolius, a member of the Calycanthaceae of magnoliids, is one of the most famous medicinal plants in Eastern China. Here, we report a chromosome‐level genome assembly of C. salicifolius, comprising 820.1 Mb of genomic sequence with a contig N50 of 2.3 Mb and containing 36 651 annotated protein‐coding genes. Phylogenetic analyses revealed that magnoliids were sister to the eudicots. Two rounds of ancient whole‐genome duplication were inferred in the C. salicifolious genome. One is shared by Calycanthaceae after its divergence with Lauraceae, and the other is in the ancestry of Magnoliales and Laurales. Notably, long genes with > 20 kb in length were much more prevalent in the magnoliid genomes compared with other angiosperms, which could be caused by the length expansion of introns inserted by transposon elements. Homologous genes within the flavonoid pathway for C. salicifolius were identified, and correlation of the gene expression and the contents of flavonoid metabolites revealed potential critical genes involved in flavonoids biosynthesis. This study not only provides an additional whole‐genome sequence from the magnoliids, but also opens the door to functional genomic research and molecular breeding of C. salicifolius.     

Genome assembly and annotation of Arabidopsis halleri,a model for heavy metal hyperaccumulation and evolutionary ecology

The self‐incompatible species Arabidopsis halleri is a close relative of the self‐compatible model plant Arabidopsis thaliana. The broad European and Asian distribution and heavy metal hyperaccumulation ability make A. halleri a useful model for ecological genomics studies. We used long‐insert mate‐pair libraries to improve the genome assembly of the A. halleri ssp. gemmifera Tada mine genotype (W302) collected from a site with high contamination by heavy metals in Japan. After five rounds of forced selfing, heterozygosity was reduced to 0.04%, which facilitated subsequent genome assembly. Our assembly now covers 196 Mb or 78% of the estimated genome size and achieved scaffold N50 length of 712 kb. To validate assembly and annotation, we used synteny of A. halleri Tada mine with a previously published high‐quality reference assembly of a closely related species, Arabidopsis lyrata. Further validation of the assembly quality comes from synteny and phylogenetic analysis of the HEAVY METAL ATPASE4 (HMA4) and METAL TOLERANCE PROTEIN1 (MTP1) regions using published sequences from European A. halleri for comparison. Three tandemly duplicated copies of HMA4, key gene involved in cadmium and zinc hyperaccumulation, were assembled on a single scaffold. The assembly will enhance the genomewide studies of A. halleri as well as the allopolyploid Arabidopsis kamchatica derived from A. lyrata and A. halleri.     

Corema album: unbiased dioecy in a competitive environment

• Corema album is a dioecious coastal shrub. Dioecious plants growing in these resource‐limited habitats may present spatial segregation of the sexes (SSS) or demographic biases because of the different reproductive effort between sexes. In these environments facilitation is a more common interaction between plants than competition. To assess factors determining the distribution of C. album male and female plants, we investigated the influence of habitat type (sand dunes and coastal woodlands), assessed the occurrence of SSS or demographic biases and also a possible role of these shrubs as nurse plants.
• We selected three C. album populations with the two habitat types. All C. album individuals were sexed, mapped and measured in three plots (20 m × 20 m) per population/habitat type. Presence and abundance of all plant species were recorded under five female and five male C. album plants as well as in equivalent open ground area in each of 15 plots.
• According to Ripley's K function result, C. album did not display SSS. Generalised linear mixed models (GLMM) show that differences in plant size were not related to plant sex. Plant inventory correspondence analysis showed that species composition and abundance were influenced by habitat type, population and the presence of a C. album individual, but not by its sex. GLMM indicated a detrimental effect of C. album on the co‐occurring plants.
• Our results show that sexual dimorphism has allowed C. album to adapt to the environment avoiding SSS or significant demographic bias, suggesting a positive outlook for its conservation.

     

Genomic variation at the tips of the adaptive radiation of Darwin's finches

Adaptive radiation unfolds as selection acts on the genetic variation underlying functional traits. The nature of this variation can be revealed by studying the tips of an ongoing adaptive radiation. We studied genomic variation at the tips of the Darwin's finch radiation; specifically focusing on polymorphism within, and variation among, three sympatric species of the genus Geospiza. Using restriction site‐associated DNA (RAD‐seq), we characterized 32 569 single‐nucleotide polymorphisms (SNPs), from which 11 outlier SNPs for beak and body size were uncovered by a genomewide association study (GWAS). Principal component analysis revealed that these 11 SNPs formed four statistically linked groups. Stepwise regression then revealed that the first PC score, which included 6 of the 11 top SNPs, explained over 80% of the variation in beak size, suggesting that selection on these traits influences multiple correlated loci. The two SNPs most strongly associated with beak size were near genes associated with beak morphology across deeper branches of the radiation: delta‐like 1 homologue (DLK1) and high‐mobility group AT‐hook 2 (HMGA2). Our results suggest that (i) key adaptive traits are associated with a small fraction of the genome (11 of 32 569 SNPs), (ii) SNPs linked to the candidate genes are dispersed throughout the genome (on several chromosomes), and (iii) micro‐ and macro‐evolutionary variation (roots and tips of the radiation) involve some shared and some unique genomic regions.     

Genome of ‘Charleston Gray’, the principal American watermelon cultivar,and genetic characterization of 1,365 accessions in the U.S. National Plant Germplasm System watermelon collection

Years of selection for desirable fruit quality traits in dessert watermelon (Citrullus lanatus) has resulted in a narrow genetic base in modern cultivars. Development of novel genomic and genetic resources offers great potential to expand genetic diversity and improve important traits in watermelon. Here, we report a high‐quality genome sequence of watermelon cultivar ‘Charleston Gray’, a principal American dessert watermelon, to complement the existing reference genome from ‘97103’, an East Asian cultivar. Comparative analyses between genomes of ‘Charleston Gray’ and ‘97103’ revealed genomic variants that may underlie phenotypic differences between the two cultivars. We then genotyped 1365 watermelon plant introduction (PI) lines maintained at the U.S. National Plant Germplasm System using genotyping‐by‐sequencing (GBS). These PI lines were collected throughout the world and belong to three Citrullus species, C. lanatus, C. mucosospermus and C. amarus. Approximately 25 000 high‐quality single nucleotide polymorphisms (SNPs) were derived from the GBS data using the ‘Charleston Gray’ genome as the reference. Population genomic analyses using these SNPs discovered a close relationship between C. lanatus and C. mucosospermus and identified four major groups in these two species correlated to their geographic locations. Citrullus amarus was found to have a distinct genetic makeup compared to C. lanatus and C. mucosospermus. The SNPs also enabled identification of genomic regions associated with important fruit quality and disease resistance traits through genome‐wide association studies. The high‐quality ‘Charleston Gray’ genome and the genotyping data of this large collection of watermelon accessions provide valuable resources for facilitating watermelon research, breeding and improvement.