Genomic variation across distribution of Micro-Tom, a model cultivar of tomato (Solanum lycopersicum).

Micro-Tom is a cultivar of tomato (Solanum lycopersicum), which is known as a major crop and model plant in Solanaceae. Micro-Tom has phenotypic traits such as dwarfism, and substantial EMS-mutagenized lines have been reported. After Micro-Tom was generated in Florida, USA, it was distributed to research institutes worldwide and used as a genetic resource. In Japan, the Micro-Tom lines have been genetically fixed; currently three lines have been re-distributed from three institutes, but many phenotypes among the lines have been observed. We have determined the genome sequence de novo of the Micro-Tom KDRI line, one of the Micro-Tom lines distributed from Kazusa DNA Research Institute (KDRI) in Japan, and have built chromosome-scale pseudomolecules. Genotypes among six Micro-Tom lines, including three in Japan, one in the United States, one in France, and one in Brazil showed phenotypic alternation. Here, we unveiled the swift emergence of genetic diversity in both phenotypes and genotypes within the Micro-Tom genome sequence during its propagation. These findings offer valuable insights crucial for the management of bioresources.

QTL analysis of femaleness in monoecious spinach and fine mapping of a major QTL using an updated version of chromosome-scale pseudomolecules.

Although spinach is predominantly dioecious, monoecious plants with varying proportions of female and male flowers are also present. Recently, monoecious inbred lines with highly female and male conditions have been preferentially used as parents for F1-hybrids, rather than dioecious lines. Accordingly, identifying the loci for monoecism is an important issue for spinach breeding. We here used long-read sequencing and Hi-C technology to construct SOL_r2.0_pseudomolecule, a set of six pseudomolecules of spinach chromosomes (total length: 879.2 Mb; BUSCO complete 97.0%) that are longer and more genetically complete than our previous version of pseudomolecules (688.0 Mb; 81.5%). Three QTLs, qFem2.1, qFem3.1, and qFem6.1, responsible for monoecism were mapped to SOL_r2.0_pseudomolecule. qFem3.1 had the highest LOD score and corresponded to the M locus, which was previously identified as a determinant of monoecious expression, by genetic analysis of progeny from female and monoecious plants. The other QTLs were shown to modulate the ratio of female to male flowers in monoecious plants harboring a dominant allele of the M gene. Our findings will enable breeders to efficiently produce highly female- and male-monoecious parental lines for F1-hybrids by pyramiding the three QTLs. Through fine-mapping, we narrowed the candidate region for the M locus to a 19.5 kb interval containing three protein-coding genes and one long non-coding RNA gene. Among them, only RADIALIS-like-2a showed a higher expression in the reproductive organs, suggesting that it might play a role in reproductive organogenesis. However, there is no evidence that it is involved in the regulation of stamen and pistil initiation, which are directly related to the floral sex differentiation system in spinach. Given that auxin is involved in reproductive organ formation in many plant species, genes related to auxin transport/response, in addition to floral organ formation, were identified as candidates for regulators of floral sex-differentiation from qFem2.1 and qFem6.1.

Whole-genome sequence analysis of mutations in rice plants regenerated from zygotes, mature embryos, and immature embryos.

Somaclonal variation was studied by whole-genome sequencing in rice plants (Oryza sativa L., 'Nipponbare') regenerated from the zygotes, mature embryos, and immature embryos of a single mother plant. The mother plant and its seed-propagated progeny were also sequenced. A total of 338 variants of the mother plant sequence were detected in the progeny, and mean values ranged from 9.0 of the seed-propagated plants to 37.4 of regenerants from mature embryos. The natural mutation rate of 1.2 × 10-8 calculated using the variants in the seed-propagated plants was consistent with the values reported previously. The ratio of single nucleotide variants (SNVs) among the variants in the seed-propagated plants was 91.1%, which is higher than 56.1% previously reported, and not significantly different from those in the regenerants. Overall, the ratio of transitions to transversions of SNVs was lower in the regenerants as shown previously. Plants regenerated from mature embryos had significantly more variants than different progeny types. Therefore, using zygotes and immature embryos can reduce somaclonal variation during the genetic manipulation of rice.

Plant GARDEN: a portal website for cross-searching between different types of genomic and genetic resources in a wide variety of plant species.

BACKGROUND: Plant genome information is fundamental to plant research and development. Along with the increase in the number of published plant genomes, there is a need for an efficient system to retrieve various kinds of genome-related information from many plant species across plant kingdoms. Various plant databases have been developed, but no public database covers both genomic and genetic resources over a wide range of plant species. MAIN BODY: We have developed a plant genome portal site, Plant GARDEN (Genome And Resource Database Entry: https://plantgarden.jp/en/index ), to provide diverse information related to plant genomics and genetics in divergent plant species. Elasticsearch is used as a search engine, and cross-keyword search across species is available. Web-based user interfaces (WUI) for PCs and tablet computers were independently developed to make data searches more convenient. Several types of data are stored in Plant GARDEN: reference genomes, gene sequences, PCR-based DNA markers, trait-linked DNA markers identified in genetic studies, SNPs, and in/dels on publicly available sequence read archives (SRAs). The data registered in Plant GARDEN as of March 2023 included 304 assembled genome sequences, 11,331,614 gene sequences, 419,132 DNA markers, 8,225 QTLs, and 5,934 SNP lists (gvcf files). In addition, we have re-annotated all the genes registered in Plant GARDEN by using a functional annotation tool, Hayai-Annotation, to compare the orthologous relationships among genes. CONCLUSION: The aim of Plant GARDEN is to provide plant genome information for use in the fields of plant science as well as for plant-based industries, education, and other relevant areas. Therefore, we have designed a WUI that allows a diverse range of users to access such information in an easy-to-understand manner. Plant GARDEN will eventually include a wide range of plant species for which genome sequences are assembled, and thus the number of plant species in the database will continue to expand. We anticipate that Plant GARDEN will promote the understanding of genomes and gene diversity by facilitating comparisons of the registered sequences.

Genome sequencing reveals the genetic architecture of heterostyly and domestication history of common buckwheat.

Common buckwheat, Fagopyrum esculentum, is an orphan crop domesticated in southwest China that exhibits heterostylous self-incompatibility. Here we present chromosome-scale assemblies of a self-compatible F. esculentum accession and a self-compatible wild relative, Fagopyrum homotropicum, together with the resequencing of 104 wild and cultivated F. esculentum accessions. Using these genomic data, we report the roles of transposable elements and whole-genome duplications in the evolution of Fagopyrum. In addition, we show that (1) the breakdown of heterostyly occurs through the disruption of a hemizygous gene jointly regulating the style length and female compatibility and (2) southeast Tibet was involved in common buckwheat domestication. Moreover, we obtained mutants conferring the waxy phenotype for the first time in buckwheat. These findings demonstrate the utility of our F. esculentum assembly as a reference genome and promise to accelerate buckwheat research and breeding.

An improved reference genome for Trifolium subterraneum L. provides insight into molecular diversity and intra-specific phylogeny.

Subterranean clover (Trifolium subterraneum L., Ts) is a geocarpic, self-fertile annual forage legume with a compact diploid genome (n = x = 8, 544 Mb/1C). Its resilience and climate adaptivity have made it an economically important species in Mediterranean and temperate zones. Using the cultivar Daliak, we generated higher resolution sequence data, created a new genome assembly TSUd_3.0, and conducted molecular diversity analysis for copy number variant (CNV) and single-nucleotide polymorphism (SNP) among 36 cultivars. TSUd_3.0 substantively improves prior genome assemblies with new Hi-C and long-read sequence data, covering 531 Mb, containing 41,979 annotated genes and generating a 94.4% BUSCO score. Comparative genomic analysis among select members of the tribe Trifolieae indicated TSUd 3.0 corrects six assembly-error inversion/duplications and confirmed phylogenetic relationships. Its synteny with T. pratense, T. repens, Medicago truncatula and Lotus japonicus genomes were assessed, with the more distantly related T. repens and M. truncatula showing higher levels of co-linearity with Ts than between Ts and its close relative T. pratense. Resequencing of 36 cultivars discovered 7,789,537 SNPs subsequently used for genomic diversity assessment and sequence-based clustering. Heterozygosity estimates ranged from 1% to 21% within the 36 cultivars and may be influenced by admixture. Phylogenetic analysis supported subspecific genetic structure, although it indicates four or five groups, rather than the three recognized subspecies. Furthermore, there were incidences where cultivars characterized as belonging to a particular subspecies clustered with another subspecies when using genomic data. These outcomes suggest that further investigation of Ts sub-specific classification using molecular and morpho-physiological data is needed to clarify these relationships. This upgraded reference genome, complemented with comprehensive sequence diversity analysis of 36 cultivars, provides a platform for future gene functional analysis of key traits, and genome-based breeding strategies for climate adaptation and agronomic performance. Pangenome analysis, more in-depth intra-specific phylogenomic analysis using the Ts core collection, and functional genetic and genomic studies are needed to further augment knowledge of Trifolium genomes.

Genome Sequence and Analysis of Nicotiana benthamiana, the Model Plant for Interactions between Organisms.

Nicotiana benthamiana is widely used as a model plant for dicotyledonous angiosperms. In fact, the strains used in research are highly susceptible to a wide range of viruses. Accordingly, these strains are subject to plant pathology and plant-microbe interactions. In terms of plant-plant interactions, N. benthamiana is one of the plants that exhibit grafting affinity with plants from different families. Thus, N. benthamiana is a good model for plant biology and has been the subject of genome sequencing analyses for many years. However, N. benthamiana has a complex allopolyploid genome, and its previous reference genome is fragmented into 141,000 scaffolds. As a result, molecular genetic analysis is difficult to perform. To improve this effort, de novo whole-genome assembly was performed in N. benthamiana with Hifi reads, and 1,668 contigs were generated with a total length of 3.1 Gb. The 21 longest scaffolds, regarded as pseudomolecules, contained a 2.8-Gb sequence, occupying 95.6% of the assembled genome. A total of 57,583 high-confidence gene sequences were predicted. Based on a comparison of the genome structures between N. benthamiana and N. tabacum, N. benthamiana was found to have more complex chromosomal rearrangements, reflecting the age of interspecific hybridization. To verify the accuracy of the annotations, the cell wall modification genes involved in grafting were analyzed, which revealed not only the previously indeterminate untranslated region, intron and open reading frame sequences but also the genomic locations of their family genes. Owing to improved genome assembly and annotation, N. benthamiana would increasingly be more widely accessible.

The genome of Lyophyllum shimeji provides insight into the initial evolution of ectomycorrhizal fungal genomes.

Mycorrhizae are one of the most fundamental symbioses between plants and fungi, with ectomycorrhizae being the most widespread in boreal forest ecosystems. Ectomycorrhizal fungi are hypothesized to have evolved convergently from saprotrophic ancestors in several fungal clades, especially members of the subdivision Agaricomycotina. Studies on fungal genomes have identified several typical characteristics of mycorrhizal fungi, such as genome size expansion and decreases in plant cell-wall degrading enzymes (PCWDEs). However, genomic changes concerning the evolutionary transition to the ectomycorrhizal lifestyle are largely unknown. In this study, we sequenced the genome of Lyophyllum shimeji, an ectomycorrhizal fungus that is phylogenetically related to saprotrophic species and retains some saprotroph-like traits. We found that the genome of Ly. shimeji strain AT787 lacks both incremental increases in genome size and reduced numbers of PCWDEs. Our findings suggest that the previously reported common genomic traits of mycorrhizal fungi are not essential for the ectomycorrhizal lifestyle, but are a result of abolishing saprotrophic activity. Since Ly. shimeji is commercially consumed as an edible mushroom, the newly available genomic information may also impact research designed to enhance the cultivation of this mushroom.

Chromosome-scale genome assembly of Eustoma grandiflorum, the first complete genome sequence in the genus Eustoma.

Eustoma grandiflorum (Raf.) Shinn. is an annual herbaceous plant native to the southern United States, Mexico, and the Greater Antilles. It has a large flower with a variety of colors and is an important flower crop. In this study, we established a chromosome-scale de novo assembly of E. grandiflorum genome sequences by integrating four genomic and genetic approaches: (1) Pacific Biosciences (PacBio) Sequel deep sequencing, (2) error correction of the assembly by Illumina short reads, (3) scaffolding by chromatin conformation capture sequencing (Hi-C), and (4) genetic linkage maps derived from an F2 mapping population. Thirty-six pseudomolecules and 64 unplaced scaffolds were created, with a total length of 1,324.8 Mb. A total of 36,619 genes were predicted on the genome as high-confidence genes. A comparison of genome structure between E. grandiflorum and C. canephora or O. pumila suggested whole-genome duplication after the divergence between the families Gentianaceae and Rubiaceae. Phylogenetic analysis with single-copy genes suggested that the divergence time between Gentianaceae and Rubiaceae was 74.94 MYA. Genetic diversity analysis was performed for nine commercial E. grandiflorum varieties bred in Japan, from which 254,205 variants were identified. This first report on the construction of a reference genome sequence in the genus Eustoma is expected to contribute to genetic and genomic studies in this genus and in the family Gentianaceae.

Genome assembly and transcriptomic analyses of the repeatedly rejuvenating jellyfish Turritopsis dohrnii.

Only two hydromedusan species, Turritopsis dohrnii and T. sp., have exhibited experimental multiple-repeat life cycle reversion in the laboratory, which can be artificially induced by various means such as incubation with CsCl, heat shock, and mechanical damage with needles. In the present study, we constructed a genome assembly of T. dohrnii using Pacific Biosciences long-reads and Illumina short-reads, for which the genome DNA was extracted from 1,500 young medusae originated from a single clone. The total length of the draft genome sequence of T. dohrnii was 435.9 Mb (N50 length 747.2 kb). We identified 23,314 high-confidence genes and found the characteristics of RNA expression amongst developmental stages. Our genome assembly and transcriptome data provide a key model system resource that will be useful for understanding cyclical rejuvenation.

Transcriptome analysis of tomato plants following salicylic acid-induced immunity against Clavibacter michiganensis ssp. michiganensis.

Salicylic acid (SA) is known to be involved in the immunity against Clavibacter michiganensis ssp. michiganensis (Cmm) that causes bacterial canker in tomato. To identify the candidate genes associated with SA-inducible Cmm resistance, transcriptome analysis was conducted via RNA sequencing in tomato plants treated with SA. SA treatment upregulated various defense-associated genes, such as PR and GST genes, in tomato cotyledons. A comparison of SA- and Cmm-responsive genes revealed that both SA treatment and Cmm infection commonly upregulated a large number of genes. Gene Ontology (GO) analysis indicated that the GO terms associated with plant immunity were over-represented in both SA- and Cmm-induced genes. The genes commonly downregulated by both SA treatment and Cmm infection were associated with the cell cycle and may be involved in growth and immunity trade-off through cell division. After SA treatment, several proteins that were predicted to play a role in immune signaling, such as resistance gene analogs, Ca2+ sensors, and WRKY transcription factors, were transcriptionally upregulated. The W-box element, which was targeted by WRKYs, was over-represented in the promoter regions of genes upregulated by both SA treatment and Cmm infection, supporting the speculation that WRKYs are important for the SA-mediated immunity against Cmm. Prediction of protein-protein interactions suggested that genes encoding receptor-like kinases and EF-hand proteins play an important role in immune signaling. Thus, various candidate genes involved in SA-inducible Cmm resistance were identified.

Chromosome-scale genome assembly of Glycyrrhiza uralensis revealed metabolic gene cluster centred specialized metabolites biosynthesis.

A high-quality genome assembly is imperative to explore the evolutionary basis of characteristic attributes that define chemotype and provide essential resources for a molecular breeding strategy for enhanced production of medicinal metabolites. Here, using single-molecule high-fidelity (HiFi) sequencing reads, we report chromosome-scale genome assembly for Chinese licorice (Glycyrrhiza uralensis), a widely used herbal and natural medicine. The entire genome assembly was achieved in eight chromosomes, with contig and scaffold N50 as 36.02 and 60.2 Mb, respectively. With only 17 assembly gaps and half of the chromosomes having no or one assembly gap, the presented genome assembly is among the best plant genomes to date. Our results showed an advantage of using highly accurate long-read HiFi sequencing data for assembling a highly heterozygous genome including its complexed repeat content. Additionally, our analysis revealed that G. uralensis experienced a recent whole-genome duplication at approximately 59.02 million years ago post a gamma (γ) whole-genome triplication event, which contributed to its present chemotype features. The metabolic gene cluster analysis identified 355 gene clusters, which included the entire biosynthesis pathway of glycyrrhizin. The genome assembly and its annotations provide an essential resource for licorice improvement through molecular breeding and the discovery of valuable genes for engineering bioactive components and understanding the evolution of specialized metabolites biosynthesis.

A genome-wide association and fine-mapping study of white rust resistance in hexaploid chrysanthemum cultivars with a wild diploid reference genome.

White rust caused by Puccinia horiana is one of the most serious diseases of chrysanthemum (Chrysanthemum × morifolium). In this study, we report the DNA markers associated with resistance against P. horiana via a simple approach using the genome of a wild diploid relative, Chrysanthemum seticuspe. First, we identified the important region of the genome in the resistant cultivar "Ariesu" via a genome-wide association study. Simplex single nucleotide polymorphism (SNP) markers mined from ddRAD-Seq were used in a biparental population originating from crosses between resistant "Ariesu" and susceptible "Yellow Queen". The C. seticuspe genome was used as a reference. For the fine mapping of P. horiana resistance locus 2 (Phr2), a comparative whole genome sequencing study was conducted. Although the genome sequences of chrysanthemum cultivars assembled via the short-read approach were fragmented, reliable genome alignments were reconstructed by mapping onto the chromosome level of the C. seticuspe pseudomolecule. Base variants were then identified by comparing the assembled genome sequences of resistant "Ariesu" and susceptible "Yellow Queen". Consequently, SNP markers that were closer to Phr2 compared with ddRAD-Seq markers were obtained. These SNP markers co-segregated with resistance in F1 progenies originating from resistant "Ariesu" and showed robust transferability for detecting Phr2-conferring resistance among chrysanthemum genetic resources. The wild C. seticuspe pseudomolecule, a de facto monoploid genome used for ddRAD-Seq analysis and assembled genome sequence comparison, demonstrated this method's utility as a model for developing DNA markers in hexaploid chrysanthemum cultivars.

De novo whole-genome assembly in an interspecific hybrid table grape, 'Shine Muscat'.

The first genome sequence of an interspecific grape hybrid (Vitis labruscana × Vitis vinifera), 'Shine Muscat', an elite table grape cultivar bred in Japan, is presented. The resultant genome assemblies included two types of sequences: a haplotype-phased sequence of the highly heterozygous genomes and an unphased sequence representing a 'pseudo-haploid' genome. The unphased sequences, assembled to the chromosome level with Hi-C reads, spanned 488.97 Mb in length, 99.1% of the estimated genome size, with 4,595 scaffold sequences and a 23.9-Mb N50 length. The phased sequences had 15,650 scaffolds spanning 1.0 Gb and a 4.2-Mb N50 length. 32,827 high-confidence genes were predicted on the unphased genomes. Clustering analysis of the 'Shine Muscat' gene sequences with three other Vitis species and Arabidopsis indicated that 11,279 orthologous gene clusters were common to Vitis spp. and Arabidopsis, 4,385 were Vitis specific, and 234 were 'Shine Muscat' specific. Whole-genome resequencing was also performed for the parental lines of 'Shine Muscat', Akitsu-21 and 'Hakunan', and parental-specific copy number variations were identified. The obtained genome resources provide new insights that could assist in cultivation and breeding strategies to produce high-quality table grapes.

Comprehensive collection of genes and comparative analysis of full-length transcriptome sequences from Japanese larch (Larix kaempferi) and Kuril larch (Larix gmelinii var. japonica).

BACKGROUND: Japanese larch (Larix kaempferi) is an economically important deciduous conifer species that grows in cool-temperate forests and is endemic to Japan. Kuril larch (L. gmelinii var. japonica) is a variety of Dahurian larch that is naturally distributed in the Kuril Islands and Sakhalin. The hybrid larch (L. gmelinii var. japonica × L. kaempferi) exhibits heterosis, which manifests as rapid juvenile growth and high resistance to vole grazing. Since these superior characteristics have been valued by forestry managers, the hybrid larch is one of the most important plantation species in Hokkaido. To accelerate molecular breeding in these species, we collected and compared full-length cDNA isoforms (Iso-Seq) and RNA-Seq short-read, and merged them to construct candidate gene as reference for both Larix species. To validate the results, candidate protein-coding genes (ORFs) related to some flowering signal-related genes ​were screened from the reference sequences, and the phylogenetic relationship with closely related species was elucidated. RESULTS: Using the isoform sequencing of PacBio RS ll and the de novo assembly of RNA-Seq short-read sequences, we identified 50,690 and 38,684 ORFs in Japanese larch and Kuril larch, respectively. BUSCO completeness values were 90.5% and 92.1% in the Japanese and Kuril larches, respectively. After comparing the collected ORFs from the two larch species, a total of 19,813 clusters, comprising 22,571 Japanese larch ORFs and 22,667 Kuril larch ORFs, were contained in the intersection of the Venn diagram. In addition, we screened several ORFs related to flowering signals (SUPPRESSER OF OVEREXPRESSION OF CO1: SOC1, LEAFY: LFY, FLOWERING Locus T: FT, CONSTANCE: CO) from both reference sequences, and very similar found in other species. CONCLUSIONS: The collected ORFs will be useful as reference sequences for molecular breeding of Japanese and Kuril larches, and also for clarifying the evolution of the conifer genome and investigating functional genomics.

Development of a genome-wide marker design workflow for onions and its application in target amplicon sequencing-based genotyping.

Onions are one of the most widely cultivated vegetables worldwide; however, the development and utilization of molecular markers have been limited because of the large genome of this plant. We present a genome-wide marker design workflow for onions and its application in a high-throughput genotyping method based on target amplicon sequencing. The efficiency of the method was evaluated by genotyping of F2 populations. In the marker design workflow, unigene and genomic sequence data sets were constructed, and polymorphisms between parental lines were detected through transcriptome sequence analysis. The positions of polymorphisms detected in the unigenes were mapped onto the genome sequence, and primer sets were designed. In total, 480 markers covering the whole genome were selected. By genotyping an F2 population, 329 polymorphic sites were obtained from the estimated positions or the flanking sequences. However, missing or sparse marker regions were observed in the resulting genetic linkage map. We modified the markers to cover these regions by genotyping the other F2 populations. The grouping and order of markers on the linkages were similar across the genetic maps. Our marker design workflow and target amplicon sequencing are useful for genome-wide genotyping of onions owing to their reliability, cost effectiveness, and flexibility.

Widespread and transgenerational retrotransposon activation in inter- and intraspecies recombinant inbred populations of Lotus japonicus.

Transposable elements (TEs) constitute a large proportion of genomes of multicellular eukaryotes, including flowering plants. TEs are normally maintained in a silenced state and their transpositions rarely occur. Hybridization between distant species has been regarded as a 'shock' that stimulates genome reorganization, including TE mobilization. However, whether crosses between genetically close parents that result in viable and fertile offspring can induce TE transpositions has remained unclear. Here, we investigated the activation of long terminal repeat (LTR) retrotransposons in three Lotus japonicus recombinant inbred line (RIL) populations. We found that at least six LTR retrotransposon families were activated and transposed in 78% of the RILs investigated. LORE1a, one of the transposed LTR retrotransposons, showed transgenerational epigenetic activation, indicating the long-term effects of epigenetic instability induced by hybridization. Our study highlights TE activation as an unexpectedly common event in plant reproduction.

Genome features of common vetch (Vicia sativa) in natural habitats.

Wild plants are often tolerant to biotic and abiotic stresses in their natural environments, whereas domesticated plants such as crops frequently lack such resilience. This difference is thought to be due to the high levels of genome heterozygosity in wild plant populations and the low levels of heterozygosity in domesticated crop species. In this study, common vetch (Vicia sativa) was used as a model to examine this hypothesis. The common vetch genome (2n = 14) was estimated as 1.8 Gb in size. Genome sequencing produced a reference assembly that spanned 1.5 Gb, from which 31,146 genes were predicted. Using this sequence as a reference, 24,118 single nucleotide polymorphisms were discovered in 1243 plants from 12 natural common vetch populations in Japan. Common vetch genomes exhibited high heterozygosity at the population level, with lower levels of heterozygosity observed at specific genome regions. Such patterns of heterozygosity are thought to be essential for adaptation to different environments. The resources generated in this study will provide insights into de novo domestication of wild plants and agricultural enhancement.

Transcriptome analysis of Clavibacter michiganensis subsp. michiganensis-infected tomatoes: a role of salicylic acid in the host response.

Bacterial canker of tomato (Solanum lycopersicon) caused by the Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis (Cmm) is an economically important disease. To understand the host defense response to Cmm infection, transcriptome sequences in tomato cotyledons were analyzed by RNA-seq. Overall, 1788 and 540 genes were upregulated and downregulated upon infection, respectively. Gene Ontology enrichment analysis revealed that genes involved in the defense response, phosphorylation, and hormone signaling were over-represented by the infection. Induced expression of defense-associated genes suggested that the tomato response to Cmm showed similarities to common plant disease responses. After infection, many resistance gene analogs (RGAs) were transcriptionally upregulated, including the expressions of some receptor-like kinases (RLKs) involved in pattern-triggered immunity. The expressions of WRKYs, NACs, HSFs, and CBP60s encoding transcription factors (TFs) reported to regulate defense-associated genes were induced after infection with Cmm. Tomato genes orthologous to Arabidopsis EDS1, EDS5/SID1, and PAD4/EDS9, which are causal genes of salicylic acid (SA)-deficient mutants, were upregulated after infection with Cmm. Furthermore, Cmm infection drastically stimulated SA accumulation in tomato cotyledons. Genes involved in the phenylalanine ammonia lyase pathway were upregulated, whereas metabolic enzyme gene expression in the isochorismate synthase pathway remained unchanged. Exogenously applied SA suppressed bacterial growth and induced the expression of WRKYs, suggesting that some Cmm-responsive genes are regulated by SA signaling, and SA signaling activation should improve tomato immunity against Cmm.

A chromosome-level genome sequence of Chrysanthemum seticuspe, a model species for hexaploid cultivated chrysanthemum.

Chrysanthemums are one of the most industrially important cut flowers worldwide. However, their segmental allopolyploidy and self-incompatibility have prevented the application of genetic analysis and modern breeding strategies. We thus developed a model strain, Gojo-0 (Chrysanthemum seticuspe), which is a diploid and self-compatible pure line. Here, we present the 3.05 Gb chromosome-level reference genome sequence, which covered 97% of the C. seticuspe genome. The genome contained more than 80% interspersed repeats, of which retrotransposons accounted for 72%. We identified recent segmental duplication and retrotransposon expansion in C. seticuspe, contributing to arelatively large genome size. Furthermore, we identified a retrotransposon family, SbdRT, which was enriched in gene-dense genome regions and had experienced a very recent transposition burst. We also demonstrated that the chromosome-level genome sequence facilitates positional cloning in C. seticuspe. The genome sequence obtained here can greatly contribute as a reference for chrysanthemum in front-line breeding including genome editing.