Tomato Genomic Resources Database: An Integrated Repository of Useful Tomato Genomic Information for Basic and Applied Research

Tomato Genomic Resources Database (TGRD) allows interactive browsing of tomato genes, micro RNAs, simple sequence repeats (SSRs), important quantitative trait loci and Tomato-EXPEN 2000 genetic map altogether or separately along twelve chromosomes of tomato in a single window. The database is created using sequence of the cultivar Heinz 1706. High quality single nucleotide polymorphic (SNP) sites between the genes of Heinz 1706 and the wild tomato S. pimpinellifolium LA1589 are also included. Genes are classified into different families. 5′-upstream sequences (5′-US) of all the genes and their tissue-specific expression profiles are provided. Sequences of the microRNA loci and their putative target genes are catalogued. Genes and 5′-US show presence of SSRs and SNPs. SSRs located in the genomic, genic and 5′-US can be analysed separately for the presence of any particular motif. Primer sequences for all the SSRs and flanking sequences for all the genic SNPs have been provided. TGRD is a user-friendly web-accessible relational database and uses CMAP viewer for graphical scanning of all the features. Integration and graphical presentation of important genomic information will facilitate better and easier use of tomato genome. TGRD can be accessed as an open source repository at http://59.163.192.91/tomato2/.     

High resolution synteny maps allowing direct comparisons between the coffee and tomato genomes

Tomato (Solanum lycopersicum) and coffee (Coffea canephora) belong to the sister families Solanaceae and Rubiaceae, respectively. We report herein the mapping of a common set of 257 Conserved Ortholog Set II genes in the genomes of both species. The mapped markers are well distributed across both genomes allowing the first syntenic comparison between species from these two families. The majority (75%) of the synteny blocks are short (<4 cM); however, some extend up to 50 cM. In an effort to further characterize the synteny between these two genomes, we took advantage of the available sequence for the tomato genome to show that tomato chromosome 7 is syntenic to half of the two coffee linkage groups E and F with the putative break point in tomato localized to the boundary of the heterochromatin and euchromatin on the long arm. In addition to the new insight on genome conservation and evolution between the plant families Solanaceae and Rubiaceae, the comparative maps presented herein provide a translational tool by which coffee researchers may take benefit of DNA sequence and genetic information from tomato and vice versa. It is thus expected that these comparative genome information will help to facilitate and expedite genetic and genomic research in coffee.     

Short interspersed nuclear elements (SINEs) are abundant in Solanaceae and have a family‐specific impact on gene structure and genome organization

Short interspersed nuclear elements (SINEs) are highly abundant non‐autonomous retrotransposons that are widespread in plants. They are short in size, non‐coding, show high sequence diversity, and are therefore mostly not or not correctly annotated in plant genome sequences. Hence, comparative studies on genomic SINE populations are rare. To explore the structural organization and impact of SINEs, we comparatively investigated the genome sequences of the Solanaceae species potato (Solanum tuberosum), tomato (Solanum lycopersicum), wild tomato (Solanum pennellii), and two pepper cultivars (Capsicum annuum). Based on 8.5 Gbp sequence data, we annotated 82 983 SINE copies belonging to 10 families and subfamilies on a base pair level. Solanaceae SINEs are dispersed over all chromosomes with enrichments in distal regions. Depending on the genome assemblies and gene predictions, 30% of all SINE copies are associated with genes, particularly frequent in introns and untranslated regions (UTRs). The close association with genes is family specific. More than 10% of all genes annotated in the Solanaceae species investigated contain at least one SINE insertion, and we found genes harbouring up to 16 SINE copies. We demonstrate the involvement of SINEs in gene and genome evolution including the donation of splice sites, start and stop codons and exons to genes, enlargement of introns and UTRs, generation of tandem‐like duplications and transduction of adjacent sequence regions.     

A COSII genetic map of the pepper genome provides a detailed picture of synteny with tomato and new insights into recent chromosome evolution in the genus Capsicum

We report herein the development of a pepper genetic linkage map which comprises 299 orthologous markers between the pepper and tomato genomes (including 263 conserved ortholog set II or COSII markers). The expected position of additional 288 COSII markers was inferred in the pepper map via pepper–tomato synteny, bringing the total orthologous markers in the pepper genome to 587. While pepper maps have been previously reported, this is the first complete map in the sense that all markers could be placed in 12 linkage groups corresponding to the 12 chromosomes. The map presented herein is relevant to the genomes of cultivated C. annuum and wild C. annuum (as well as related Capsicum species) which differ by a reciprocal chromosome translocation. This map is also unique in that it is largely based on COSII markers, which permits the inference of a detailed syntenic relationship between the pepper and tomato genomes—shedding new light on chromosome evolution in the Solanaceae. Since divergence from their last common ancestor is approximately 20 million years ago, the two genomes have become differentiated by a minimum number of 19 inversions and 6 chromosome translocations, as well as numerous putative single gene transpositions. Nevertheless, the two genomes share 35 conserved syntenic segments (CSSs) within which gene/marker order is well preserved. The high resolution COSII synteny map described herein provides a platform for cross-reference of genetic and genomic information (including the tomato genome sequence) between pepper and tomato and therefore will facilitate both applied and basic research in pepper. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Cross-species bacterial artificial chromosome-fluorescence in situ hybridization painting of the tomato and potato chromosome 6 reveals undescribed chromosomal rearrangements

Ongoing genomics projects of tomato (Solanum lycopersicum) and potato (S. tuberosum) are providing unique tools for comparative mapping studies in Solanaceae. At the chromosomal level, bacterial artificial chromosomes (BACs) can be positioned on pachytene complements by fluorescence in situ hybridization (FISH) on homeologous chromosomes of related species. Here we present results of such a cross-species multicolor cytogenetic mapping of tomato BACs on potato chromosomes 6 and vice versa. The experiments were performed under low hybridization stringency, while blocking with Cot-100 was essential in suppressing excessive hybridization of repeat signals in both within-species FISH and cross-species FISH of tomato BACs. In the short arm we detected a large paracentric inversion that covers the whole euchromatin part with breakpoints close to the telomeric heterochromatin and at the border of the short arm pericentromere. The long arm BACs revealed no deviation in the colinearity between tomato and potato. Further comparison between tomato cultivars Cherry VFNT and Heinz 1706 revealed colinearity of the tested tomato BACs, whereas one of the six potato clones (RH98-856-18) showed minor putative rearrangements within the inversion. Our results present cross-species multicolor BAC–FISH as a unique tool for comparative genetic studies across Solanum species.     

Fine mapping of the Ph-3 gene conferring resistance to late blight (Phytophthora infestans) in tomato

Late blight, caused by the oomycete pathogen Phytophthora infestans (Mont.) de Bary, is a devastating disease for tomato and potato crops. In the past decades, many late blight resistance (R) genes have been characterized in potato. In contrast, less work has been conducted on tomato. The Ph-3 gene from Solanum pimpinellifolium was introgressed into cultivated tomatoes and conferred broad-spectrum resistance to P. infestans. It was previously assigned to the long arm of chromosome 9. In this study, a high-resolution genetic map covering the Ph-3 locus was constructed using an F₂ population of a cross between Solanum lycopersicum CLN2037B (containing Ph-3) and S. lycopersicum LA4084. Ph-3 was mapped in a 0.5 cM interval between two markers, Indel_3 and P55. Eight putative genes were found in the corresponding 74 kb region of the tomato Heinz1706 reference genome. Four of these genes are resistance gene analogs (RGAs) with a typical nucleotide-binding adaptor shared by APAF-1, R proteins, and CED-4 domain. Each RGA showed high homology to the late blight R gene Rpi-vnt1.1 from Solanum venturii. Transient gene silencing indicated that a member of this RGA family is required for Ph-3-mediated resistance to late blight in tomato. Furthermore, this RGA family was also found in the potato genome, but the number of the RGAs was higher than in tomato.     

Extensive structural variations between mitochondrial genomes of CMS and normal peppers (Capsicum annuum L.) revealed by complete nucleotide sequencing

Background

Cytoplasmic male sterility (CMS) is an inability to produce functional pollen that is caused by mutation of the mitochondrial genome. Comparative analyses of mitochondrial genomes of lines with and without CMS in several species have revealed structural differences between genomes, including extensive rearrangements caused by recombination. However, the mitochondrial genome structure and the DNA rearrangements that may be related to CMS have not been characterized in Capsicum spp.

Results

We obtained the complete mitochondrial genome sequences of the pepper CMS line FS4401 (507,452 bp) and the fertile line Jeju (511,530 bp). Comparative analysis between mitochondrial genomes of peppers and tobacco that are included in Solanaceae revealed extensive DNA rearrangements and poor conservation in non-coding DNA. In comparison between pepper lines, FS4401 and Jeju mitochondrial DNAs contained the same complement of protein coding genes except for one additional copy of an atp6 gene (ψatp6-2) in FS4401. In terms of genome structure, we found eighteen syntenic blocks in the two mitochondrial genomes, which have been rearranged in each genome. By contrast, sequences between syntenic blocks, which were specific to each line, accounted for 30,380 and 17,847 bp in FS4401 and Jeju, respectively. The previously-reported CMS candidate genes, orf507 and ψatp6-2, were located on the edges of the largest sequence segments that were specific to FS4401. In this region, large number of small sequence segments which were absent or found on different locations in Jeju mitochondrial genome were combined together. The incorporation of repeats and overlapping of connected sequence segments by a few nucleotides implied that extensive rearrangements by homologous recombination might be involved in evolution of this region. Further analysis using mtDNA pairs from other plant species revealed common features of DNA regions around CMS-associated genes.

Conclusions

Although large portion of sequence context was shared by mitochondrial genomes of CMS and male-fertile pepper lines, extensive genome rearrangements were detected. CMS candidate genes located on the edges of highly-rearranged CMS-specific DNA regions and near to repeat sequences. These characteristics were detected among CMS-associated genes in other species, implying a common mechanism might be involved in the evolution of CMS-associated genes.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-561) contains supplementary material, which is available to authorized users.     

Comprehensive Resources for Tomato Functional Genomics Based on the Miniature Model Tomato Micro-Tom

Tomato (Solanum lycopersicum L., Solanaceae) is an excellent model plant for genomic research of solanaceous plants, as well as for studying the development, ripening, and metabolism of fruit. In 2003, the International Solanaceae Project (SOL, www.sgn.cornell.edu ) was initiated by members from more than 30 countries, and the tomato genome-sequencing project is currently underway. Genome sequence of tomato obtained by this project will provide a firm foundation for forthcoming genomic studies such as the comparative analysis of genes conserved among the Solanaceae species and the elucidation of the functions of unknown tomato genes. To exploit the wealth of the genome sequence information, there is an urgent need for novel resources and analytical tools for tomato functional genomics. Here, we present an overview of the development of genetic and genomic resources of tomato in the last decade, with a special focus on the activities of Japan SOL and the National Bio-Resource Project in the development of functional genomic resources of a model cultivar, Micro-Tom.Key Words: Expressed sequence tag, full-length cDNA, genome sequence, mutant resource, Micro-Tom, JSOL, Solanum lycopersicum, tomato.     

Draft Genome Sequence of Eggplant (Solanum melongena L.): the Representative Solanum Species Indigenous to the Old World

Unlike other important Solanaceae crops such as tomato, potato, chili pepper, and tobacco, all of which originated in South America and are cultivated worldwide, eggplant (Solanum melongena L.) is indigenous to the Old World and in this respect it is phylogenetically unique. To broaden our knowledge of the genomic nature of solanaceous plants further, we dissected the eggplant genome and built a draft genome dataset with 33,873 scaffolds termed SME_r2.5.1 that covers 833.1 Mb, ca. 74% of the eggplant genome. Approximately 90% of the gene space was estimated to be covered by SME_r2.5.1 and 85,446 genes were predicted in the genome. Clustering analysis of the predicted genes of eggplant along with the genes of three other solanaceous plants as well as Arabidopsis thaliana revealed that, of the 35,000 clusters generated, 4,018 were exclusively composed of eggplant genes that would perhaps confer eggplant-specific traits. Between eggplant and tomato, 16,573 pairs of genes were deduced to be orthologous, and 9,489 eggplant scaffolds could be mapped onto the tomato genome. Furthermore, 56 conserved synteny blocks were identified between the two species. The detailed comparative analysis of the eggplant and tomato genomes will facilitate our understanding of the genomic architecture of solanaceous plants, which will contribute to cultivation and further utilization of these crops.     

Resistance gene enrichment sequencing (RenSeq) enables reannotation of the NB‐LRR gene family from sequenced plant genomes and rapid mapping of resistance loci in segregating populations

RenSeq is a NB‐LRR (nucleotide binding‐site leucine‐rich repeat) gene‐targeted, Resistance gene enrichment and sequencing method that enables discovery and annotation of pathogen resistance gene family members in plant genome sequences. We successfully applied RenSeq to the sequenced potato Solanum tuberosum clone DM, and increased the number of identified NB‐LRRs from 438 to 755. The majority of these identified R gene loci reside in poorly or previously unannotated regions of the genome. Sequence and positional details on the 12 chromosomes have been established for 704 NB‐LRRs and can be accessed through a genome browser that we provide. We compared these NB‐LRR genes and the corresponding oligonucleotide baits with the highest sequence similarity and demonstrated that ~80% sequence identity is sufficient for enrichment. Analysis of the sequenced tomato S. lycopersicum ‘Heinz 1706’ extended the NB‐LRR complement to 394 loci. We further describe a methodology that applies RenSeq to rapidly identify molecular markers that co‐segregate with a pathogen resistance trait of interest. In two independent segregating populations involving the wild Solanum species S. berthaultii (Rpi‐ber2) and S. ruiz‐ceballosii (Rpi‐rzc1), we were able to apply RenSeq successfully to identify markers that co‐segregate with resistance towards the late blight pathogen Phytophthora infestans. These SNP identification workflows were designed as easy‐to‐adapt Galaxy pipelines.     

In silico prediction of proteins related to xyloglucan fucosyltransferases in Solanaceae genomes

Two independent studies have shown that the cell wall of pollen tubes from tobacco and tomato species contained fucosylated xyloglucan (XyG). These findings are intriguing as many reports have shown that XyG of somatic cells of these species is not fucosylated but instead is arabinosylated. In order to produce fucosylated XyG, plants must express a functional galactoside α-2-fucosyltransferase. Here, using a bioinformatics approach, we show that several candidate genes coding for XyG fucosyltransferases are present in the genome of coffee and several Solanaceae species including tomato, tobacco, potato, eggplant and pepper. BLAST and protein alignments with the 2 well-characterized XyG fucosyltransferases from Arabidopsis thaliana and Pisum sativum revealed that at least 6 proteins from different Solanaceae species and from coffee displayed the 3 conserved motifs required for XyG fucosyltransferase activity.     

Complete sequencing and comparative analyses of the pepper (Capsicum annuum L.) plastome revealed high frequency of tandem repeats and large insertion/deletions on pepper plastome

Plants in the family Solanaceae are used as model systems in comparative and evolutionary genomics. The complete chloroplast genomes of seven solanaceous species have been sequenced, including tobacco, potato and tomato, but not peppers. We analyzed the complete chloroplast genome sequence of the hot pepper, Capsicum annuum. The pepper chloroplast genome was 156,781 bp in length, including a pair of inverted repeats (IR) of 25,783 bp. The content and the order of 133 genes in the pepper chloroplast genome were identical to those of other solanaceous plastomes. To characterize pepper plastome sequence, we performed comparative analysis using complete plastome sequences of pepper and seven solanaceous plastomes. Frequency and contents of large indels and tandem repeat sequences and distribution pattern of genome-wide sequence variations were investigated. In addition, a phylogenetic analysis using concatenated alignments of coding sequences was performed to determine evolutionary position of pepper in Solanaceae. Our results revealed two distinct features of pepper plastome compared to other solanaceous plastomes. Firstly, large indels, including insertions on accD and rpl20 gene sequences, were predominantly detected in the pepper plastome compared to other solanaceous plastomes. Secondly, tandem repeat sequences were particularly frequent in the pepper plastome. Taken together, our study represents unique features of evolution of pepper plastome among solanaceous plastomes.     

Identification, Phylogeny, and Expression Analysis of Pto-like Genes in Pepper

The Pto gene from the wild tomato (Solanum pimpinellifolium Mill.) encodes a serine/threonine kinase that plays an important role in bacterial speck resistance in the cultivated tomato (Solanum lycopersicum Mill.). In this paper, 10 classes of Pto-like genes are identified using degenerate polymerase chain reaction (PCR) primers and database mining in pepper. Sequences alignment reveals that many features of the gene family, such as subdomains, autophosphorylation sites, and important amino acid residues for tomato Pto, are well conserved in pepper. A phylogenetic tree of pepper Pto-like genes along with those of other plant species, including tomato Pto genes, suggests that these genes share a common evolutionary origin, and they may have evolved prior to the divergence of monocotyledonous and dicotyledonous plants. Expression analysis has revealed that nine selected Pto-like genes can be detected in at least one of the tissues grown under normal growth conditions; however, these genes are differentially expressed. In addition, some of these genes are regulated by at least one of the subjected treatments, including hormones, abiotic stress, and pathogen infection. These findings will contribute to expanding our knowledge of the roles of Pto-like genes in growth, development, and stress tolerance in pepper.     

The Ph-3 gene from Solanum pimpinellifolium encodes CC-NBS-LRR protein conferring resistance to Phytophthora infestans

Key message

Ph-3 is the first cloned tomato gene for resistance to late blight and encodes a CC-NBS-LRR protein.

Abstract

Late blight, caused by Phytophthora infestans, is one of the most destructive diseases in tomato. The resistance (R) gene Ph-3, derived from Solanum pimpinellifolium L3708, provides resistance to multiple P. infestans isolates and has been widely used in tomato breeding programmes. In our previous study, Ph-3 was mapped into a region harbouring R gene analogues (RGA) at the distal part of long arm of chromosome 9. To further narrow down the Ph-3 interval, more recombinants were identified using the flanking markers G2-4 and M8-2, which defined the Ph-3 gene to a 26 kb region according to the Heinz1706 reference genome. To clone the Ph-3 gene, a bacterial artificial chromosome (BAC) library was constructed using L3708 and one BAC clone B25E21 containing the Ph-3 region was identified. The sequence of the BAC clone B25E21 showed that only one RGA was present in the target region. A subsequent complementation analysis demonstrated that this RGA, encoding a CC-NBS-LRR protein, was able to complement the susceptible phenotype in cultivar Moneymaker. Thus this RGA was considered the Ph-3 gene. The predicted Ph-3 protein shares high amino acid identity with the chromosome-9-derived potato resistance proteins against P. infestans (Rpi proteins).     

In Silico Identification and Experimental Validation of Insertion–Deletion Polymorphisms in Tomato Genome

Comparative analysis of the genome sequences of Solanum lycopersicum variety Heinz 1706 and S. pimpinellifolium accession LA 1589 using MUGSY software identified 145 695 insertion–deletion (InDel) polymorphisms. A selected set of 3029 candidate InDels (≥2 bp) across the entire tomato genome were subjected to PCR validation, and 82.4% could be verified. Of 2272 polymorphic InDels between LA 1589 and Heinz 1706, 61.6, 45.2, and 31.6% were polymorphic in 8 accessions of S. pimpinellifolium, 4 accessions of S. lycopersicum var. cerasiforme, and 10 varieties of S. lycopersicum, respectively. Genetic distance was 0.216 in S. pimpinellifolium, 0.202 in S. lycopersicum var. cerasiforme, and 0.108 in S. lycopersicum. The data suggested a reduction of genetic variation from S. pimpinellifolium to S. lycopersicum var. cerasiforme and S. lycopersicum. Cluster analysis showed that the 8 accessions of S. pimpinellifolium were in one group, whereas 4 accessions of S. lycopersicum var. cerasiforme and 10 varieties of S. lycopersicum were in the same group.     

Phylogeny and Molecular Evolution of miR820 and miR396 microRNA Families in <Emphasis Type="Italic">Oryza</Emphasis> AA Genomes

The phylogeny and evolution of the microRNA families, miR820 and miR396, was analysed across the AA genomes of the Oryza species, the close relatives of domesticated rice. A highly dynamic evolution of the miR820 family was revealed. The number of copies of MIR820 genes, their chromosomal location and the mature microRNA sequence varied greatly with a total of 16 novel miR820 variants being identified. The phylogeny of pre-MIR820 sequences revealed that MIR820 genes of recently evolved Oryza AA genomes may have derived from sequence divergence of one or a few ancestral genes found in wild Australian perennial rice populations, Taxon B (jpn2)-MIR820 genes. Genomic scale duplication played an important role in the evolution of some of the miR396 family genes in AA genome Oryza species. miR396 family contained a MIR396 gene cluster (MIR396a and MIR396c) which was conserved across the cereal genomes. Nucleotide diversity analysis at these two MIR396 loci revealed that domesticated rice has retained less than 10% of the total diversity present in wild species. In contrast, the nucleotide sequence of four MIR396 loci remained almost conserved across domesticated and wild rices, indicating that they were under extreme functional constraint and may be involved in regulating some fundamental processes which are important both for wild and domesticated rices. Expression analysis demonstrated that miR820 variants were expressed in O. glaberrima O. barthi and O. longistaminata genome. These findings pose new challenges to explain the possible role of miR820 variants identified.