Nucleotide-binding Site Leucine-rich Repeat Research Articles

Cytological and molecular characterization of wheat lines carrying leaf rust and stem rust resistance genes Lr24 and Sr24

Previous studies showed that Australian wheat cultivars Janz and Sunco carry leaf rust and stem rust resistance genes Lr24 and Sr24 derived from Thinopyrum ponticum chromosome arm 3AgL. However, the size of the alien segments carrying Lr24 and Sr24 in the lines were not determined. In this study, we used non-denaturing fluorescence in situ hybridization (ND-FISH), genomic in situ hybridization (GISH), and PCR-based landmark unique gene (PLUG) markers to visualize the alien segments in Janz and Sunco, and further compared them with the segments in US cultivars Agent and Amigo. The fraction length (FL) of the alien translocation in Agent was 0.70–1.00, whereas those in Janz, Sunco, and Amigo were smaller, at FL 0.85–1.00. It was deduced that the alien gene RAg encoding for red grain color and rust resistance genes Lr24 and Sr24 on chromosome arm 3AgL were in bins of FL 0.70–0.85 and 0.85–1.00, respectively. We retrieved and extracted nucleotide-binding site-leucine-rich repeat (NBS-LRR) receptor genes corresponding to the region of Lr24 and Sr24 on chromosomes 3E, and 3J, 3Js and 3St from the reference genome sequences of Th. elongatum and Th. intermedium, respectively. A set of molecular markers developed for Lr24 and Sr24 from those extracted NBS-LRR genes will provide valuable information for fine mapping and cloning of these genes.

GWAS analysis reveals the genetic basis of blast resistance associated with heading date in rice.

Rice blast is a destructive fungal disease affecting rice plants at various growth stages, significantly threatening global yield stability. Development of resistant rice cultivars stands as a practical means of disease control. Generally, association mapping with a diversity panel powerfully identifies new alleles controlling trait of interest. On the other hand, utilization of a breeding panel has its advantage that can be directly applied in a breeding program. In this study, we conducted a genome-wide association study (GWAS) for blast resistance using 296 commercial rice cultivars with low population structure but large phenotypic diversity. We attempt to answer the genetic basis behind rice blast resistance among early maturing cultivars by subdividing the population based on its Heading date 1 (Hd1) functionality. Subpopulation-specific GWAS using the mixed linear model (MLM) based on blast nursery screening conducted in three years revealed a total of 26 significant signals, including three nucleotide-binding site leucine-rich repeat (NBS-LRR) genes (Os06g0286500, Os06g0286700, and Os06g0287500) located at Piz locus on chromosome 6, and one at the Pi-ta locus (Os12g0281300) on chromosome 12. Haplotype analysis revealed blast resistance associated with Piz locus was exclusively specific to Type 14 hd1 among japonica rice. Our findings provide valuable insights for breeding blast resistant rice and highlight the applicability of our elite cultivar panel to detect superior alleles associated with important agronomic traits.

Insights into the genetic architecture of Phytophthora capsici root rot resistance in chile pepper (Capsicum spp.) from multi-locus genome-wide association study

BackgroundPhytophthora root rot, a major constraint in chile pepper production worldwide, is caused by the soil-borne oomycete, Phytophthora capsici. This study aimed to detect significant regions in the Capsicum genome linked to Phytophthora root rot resistance using a panel consisting of 157 Capsicum spp. genotypes. Multi-locus genome wide association study (GWAS) was conducted using single nucleotide polymorphism (SNP) markers derived from genotyping-by-sequencing (GBS). Individual plants were separately inoculated with P. capsici isolates, ‘PWB-185’, ‘PWB-186’, and ‘6347’, at the 4–8 leaf stage and were scored for disease symptoms up to 14-days post-inoculation. Disease scores were used to calculate disease parameters including disease severity index percentage, percent of resistant plants, area under disease progress curve, and estimated marginal means for each genotype.ResultsMost of the genotypes displayed root rot symptoms, whereas five accessions were completely resistant to all the isolates and displayed no symptoms of infection. A total of 55,117 SNP markers derived from GBS were used to perform multi-locus GWAS which identified 330 significant SNP markers associated with disease resistance. Of these, 56 SNP markers distributed across all the 12 chromosomes were common across the isolates, indicating association with more durable resistance. Candidate genes including nucleotide-binding site leucine-rich repeat (NBS-LRR), systemic acquired resistance (SAR8.2), and receptor-like kinase (RLKs), were identified within 0.5 Mb of the associated markers.ConclusionsResults will be used to improve resistance to Phytophthora root rot in chile pepper by the development of Kompetitive allele-specific markers (KASP®) for marker validation, genomewide selection, and marker-assisted breeding.

Genome-wide characterization of the NBLRR gene family provides evolutionary and functional insights into blast resistance in pearl millet (Cenchrusamericanus (L.) Morrone).

The investigation is the first report on genome-wide identification and characterization of NBLRR genes in pearl millet. We have shown the role of gene loss and purifying selection in the divergence of NBLRRs in Poaceae lineage and candidate CaNBLRR genes for resistance to Magnaporthe grisea infection. Plants have evolved multiple integral mechanisms to counteract the pathogens' infection, among which plant immunity through NBLRR (nucleotide-binding site, leucine-rich repeat) genes is at the forefront. The genome-wide mining in pearl millet (Cenchrus americanus (L.) Morrone) revealed 146 CaNBLRRs. The variation in the branch length of NBLRRs showed the dynamic nature of NBLRRs in response to evolving pathogen races. The orthology of NBLRRs showed a predominance of many-to-one orthologs, indicating the divergence of NBLRRs in the pearl millet lineage mainly through gene loss events followed by gene gain through single-copy duplications. Further, the purifying selection (Ka/Ks < 1) shaped the expansion of NBLRRs within the lineage of pear millet and other members of Poaceae. Presence of cis-acting elements, viz. TCA element, G-box, MYB, SARE, ABRE and conserved motifs annotated with P-loop, kinase 2, RNBS-A, RNBS-D, GLPL, MHD, Rx-CC and LRR suggests their putative role in disease resistance and stress regulation. The qRT-PCR analysis in pearl millet lines showing contrasting responses to Magnaporthe grisea infection identified CaNBLRR20, CaNBLRR33, CaNBLRR46 CaNBLRR51, CaNBLRR78 and CaNBLRR146 as putative candidates. Molecular docking showed the involvement of three and two amino acid residues of LRR domains forming hydrogen bonds (histidine, arginine and threonine) and salt bridges (arginine and lysine) with effectors. Whereas 14 and 20 amino acid residues of CaNBLRR78 and CaNBLRR20 showed hydrophobic interactions with 11 and 9 amino acid residues of effectors, Mg.00g064570.m01 and Mg.00g006570.m01, respectively. The present investigation gives a comprehensive overview of CaNBLRRs and paves the foundation for their utility in pearl millet resistance breeding through understanding of host-pathogen interactions.

The soybean Rpp3 gene encodes a TIR-NBS-LRR protein that confers resistance to Phakopsora pachyrhizi.

Soybean rust is an economically significant disease caused by the fungus Phakopsora pachyrhizi that negatively impacts soybean (Glycine max (L.) Merr.) production throughout the world. Susceptible plants infected by P. pachyrhizi develop tan-colored lesions on the leaf surface that give rise to funnel-shaped uredinia as the disease progresses. While most soybean germplasm is susceptible, seven genetic loci (Rpp1 to Rpp7) that provide race-specific resistance to P. pachyrhizi (Rpp) have been identified. Rpp3 was first discovered and characterized in the soybean accession PI 462312 (Ankur), and it was also determined to be one of two Rpp genes present in PI 506764 (Hyuuga). Genetic crosses with PI 506764 were later used to fine-map the Rpp3 locus to a 371 kb region on chromosome 6. The corresponding region in the susceptible Williams 82 (Wm82) reference genome contains several homologous nucleotide binding site-leucine rich repeat (NBS-LRR) genes. To identify Rpp3, we designed oligonucleotide primers to amplify Rpp3 candidate (Rpp3C) NBS-LRR genes at this locus from PI 462312, PI 506764, and Wm82 using polymerase chain reaction (PCR). Five Rpp3C genes were identified in both Rpp3-resistant soybean lines, and co-silencing these genes compromised resistance to P. pachyrhizi. Gene expression analysis and sequence comparisons of the Rpp3C genes in PI 462312 and PI 506764 suggest that a single candidate gene, Rpp3C3, is responsible for Rpp3-mediated resistance.

Development of NBS-LRR gene-specific microsatellite markers for passion fruit Passiflora edulis Sims

The chromosome-level genome assembly of passion fruit (Passiflora edulis Sims) has facilitated computational analyses in developing novel informative SSR markers for genetics and breeding research. In this study, in-silico analysis was conducted to discover potential microsatellite loci and to design specific primers from the sequences of the characterised nucleotide-binding site - leucine-rich repeats (NBS-LRR) gene family of passion fruit, and a genotyping experiment was performed to validate the novel simple sequence repeat (SSR) markers.In total, 497 microsatellite loci were detected from 161 passion fruit NBS-LRR genes, of which 383 loci were qualified for primer generation. Forty-six SSR markers were selected for polymorphism verification, targeting 35 NBS-LRR genes that showed potential gene expression in response to biotic and abiotic stresses. From that, 25 SSR markers showed consistent polymorphic alleles among different passion fruit genotypes via PCR-PAGE analysis. These novel NBS-LRR-specific SSR markers might provide valuable tools for fortifying research on germplasm evaluation, genetic mapping, and molecular breeding for disease-resistant passion fruit varieties.

Genome-wide association study (GWAS) reveals polygenic architecture for limber pine quantitative disease resistance to white pine blister rust.

Development of durable resistance effective against a broad range of pathotypes is crucial for restoration of pathogen-damaged ecosystems. This study dissected the complex genetic architecture for limber pine quantitative disease resistance (QDR) to Cronartium ribicola using a genome-wide association study (GWAS). Eighteen-month-old seedlings were inoculated for resistance screening under controlled conditions. Disease development was quantitatively assessed for QDR-related traits over four years post inoculation. To reveal genomic architecture contributing to QDR-related traits, a set of genes related to disease resistance with genome-wide distribution was selected for targeted sequencing for genotyping of single-nucleotide polymorphisms (SNPs). GWAS revealed a set of SNPs significantly associated with quantitative traits for limber pine QDR to WPBR, including number of needle spots and stem cankers, as well as survival four years post-inoculation. The peaks of marker-trait associations displayed a polygenic pattern with genomic regions as potential resistant quantitative trait loci (QTLs), distributed over ten of the 12 linkage groups (LGs) of Pinus. None of them were linked to the Cr4-controlled major gene resistance (MGR) previously mapped on LG08. Both normal canker and bole infection were mapped on LG05, and the associated SNPs explained their phenotypic variance up to 52%, tagging a major resistant QTL. Candidate genes containing phenotypically associated SNPs encoded putative nucleotide-binding site leucine-rich repeat (NBS-LRR) proteins, LRR-receptor-like kinase (LRR-RLK), cytochrome P450 superfamily protein, heat shock cognate protein 70, glutamate receptor, RNA-binding family protein, and unknown protein. The confirmation of resistant QTLs broadens the genetic pool of limber pine resistance germplasm for resistance breeding.

Genome-wide identification and characterization of NBLRR genes in finger millet (Eleusine coracana L.) and their expression in response to Magnaporthe grisea infection

BackgroundThe nucleotide binding site leucine rich repeat (NBLRR) genes significantly regulate defences against phytopathogens in plants. The genome-wide identification and analysis of NBLRR genes have been performed in several species. However, the detailed evolution, structure, expression of NBLRRs and functional response to Magnaporthe grisea are unknown in finger millet (Eleusine coracana (L.) Gaertn.).ResultsThe genome-wide scanning of the finger millet genome resulted in 116 NBLRR (EcNBLRRs1-116) encompassing 64 CC-NB-LRR, 47 NB-LRR and 5 CCR-NB-LRR types. The evolutionary studies among the NBLRRs of five Gramineae species, viz., purple false brome (Brachypodium distachyon (L.) P.Beauv.), finger millet (E. coracana), rice (Oryza sativa L.), sorghum (Sorghum bicolor L. (Moench)) and foxtail millet (Setaria italica (L.) P.Beauv.) showed the evolution of NBLRRs in the ancestral lineage of the target species and subsequent divergence through gene-loss events. The purifying selection (Ka/Ks < 1) shaped the expansions of NBLRRs paralogs in finger millet and orthologs among the target Gramineae species. The promoter sequence analysis showed various stress- and phytohormone-responsive cis-acting elements besides growth and development, indicating their potential role in disease defence and regulatory mechanisms. The expression analysis of 22 EcNBLRRs in the genotypes showing contrasting responses to Magnaporthe grisea infection revealed four and five EcNBLRRs in early and late infection stages, respectively. The six of these nine candidate EcNBLRRs proteins, viz., EcNBLRR21, EcNBLRR26, EcNBLRR30, EcNBLRR45, EcNBLRR55 and EcNBLRR76 showed CC, NB and LRR domains, whereas the EcNBLRR23, EcNBLRR32 and EcNBLRR83 showed NB and LRR somains.ConclusionThe identification and expression analysis of EcNBLRRs showed the role of EcNBLRR genes in assigning blast resistance in finger millet. These results pave the foundation for in-depth and targeted functional analysis of EcNBLRRs through genome editing and transgenic approaches.

Uncovering the transcriptional responses of tobacco (Nicotiana tabacum L.) roots to Ralstonia solanacearum infection: a comparative study of resistant and susceptible cultivars

BackgroundTobacco bacterial wilt (TBW) caused by Ralstonia solanacearum is the most serious soil-borne disease of tobacco that significantly reduces crop yield. However, the limited availability of resistance in tobacco hinders breeding efforts for this disease.ResultsIn this study, we conducted hydroponic experiments for the root expression profiles of D101 (resistant) and Honghuadajinyuan (susceptible) cultivars in response to BW infection at 0 h, 6 h, 1 d, 3 d, and 7d to explore the defense mechanisms of BW resistance in tobacco. As a result, 20,711 and 16,663 (total: 23,568) differentially expressed genes (DEGs) were identified in the resistant and susceptible cultivars, respectively. In brief, at 6 h, 1 d, 3 d, and 7 d, the resistant cultivar showed upregulation of 1553, 1124, 2583, and 7512 genes, while the susceptible cultivar showed downregulation of 1213, 1295, 813, and 7735 genes. Similarly, across these time points, the resistant cultivar had downregulation of 1034, 749, 1686, and 11,086 genes, whereas the susceptible cultivar had upregulation of 1953, 1790, 2334, and 6380 genes. The resistant cultivar had more up-regulated genes at 3 d and 7 d than the susceptible cultivar, indicating that the resistant cultivar has a more robust defense response against the pathogen. The GO and KEGG enrichment analysis showed that these genes are involved in responses to oxidative stress, plant–pathogen interactions, cell walls, glutathione and phenylalanine metabolism, and plant hormone signal transduction. Among the DEGs, 239 potential candidate genes were detected, including 49 phenylpropane/flavonoids pathway-associated, 45 glutathione metabolic pathway-associated, 47 WRKY, 48 ERFs, eight ARFs, 26 pathogenesis-related genes (PRs), and 14 short-chain dehydrogenase/reductase genes. In addition, two highly expressed novel genes (MSTRG.61386-R1B-17 and MSTRG.61568) encoding nucleotide-binding site leucine-rich repeat (NBS-LRR) proteins were identified in both cultivars at 7 d.ConclusionsThis study revealed significant enrichment of DEGs in GO and KEGG terms linked to glutathione, flavonoids, and phenylpropane pathways, indicating the potential role of glutathione and flavonoids in early BW resistance in tobacco roots. These findings offer fundamental insight for further exploration of the genetic architecture and molecular mechanisms of BW resistance in tobacco and solanaceous plants at the molecular level.

IN-SILICO DIVERSITY ANALYSIS OF DISEASE RESISTANT NBS-LRR PROTEIN VARIANTS IN ORYZA SATIVA

Objective: Oryza sativa being the contributor of more than 2 million tons of staple food plays assorted role in agrarian economy of Pakistan. Variety of pathogens and insecticides cause serious loses of its yield. To develop immunity at genetic level in O. sativa, nucleotide binding site-leucine rich repeat proteins (NBS-LRR), associated with plant response to pathogens, have been targeted in present study.Methodology: Eight nucleotide sequences of NBS-LRR protein were retrieved from Rice Genome Annotation Project (RGAP) database. Variants were analyzed through ProtParam, CELLO2 and SOPMA tools and SWISS MODEL server.Results and conclusion: Isoelectric point (pI), instability index, aliphatic index and GRAVY showed only slight variations. Diversity was observed in sub-cellular localization, secondary (2D) and tertiary (3D) structures. Significant heterogeneity found among NBS-LRR proteins, in present study, might help to remove homogeneity in rice crops leading to reduced epidemics and yield stability.

Soybean Variety Saedanbaek Confers a New Resistance Allele to Phytophthora sojae

Phytophthora root and stem rot (PRSR) disease results in substantial losses in soybean production worldwide. The occurrence of PRSR caused by Phytophthora sojae Kaufmann &amp; Gerdemann has become increasingly important for soybean production in the Republic of Korea, but domestic soybean–P. sojae interaction has been less studied. The disease has been managed by developing varieties harboring resistance to the Phytophthora sojae (Rps) gene. The present study aimed to identify a major gene locus conferring resistance to new P. sojae isolate 2858 in the recombinant inbred line population derived from a cross between parental lines ‘Daepung’ (susceptible) and ‘Saedanbaek’ (resistant). Seventy-three recombination inbred lines (RILs) were evaluated for resistance to P. sojae isolate 2858. A resistance locus was identified in the approximate 3.3–4.3 megabase pair region on chromosome 3 using both single-marker and linkage analyses. The Rps of Saedanbaek (RpsSDB) was located on the well-known Rps gene/allele cluster region, which also partially overlapped with a locus previously identified in the Korean soybean variety, ‘Daewon’, resistant to another P. sojae isolate 2457 (RpsDW). Approximately 402 kilobase pairs of the interval region overlapped, including six nucleotide-binding site-leucine-rich repeat (NBS-LRR)-coding genes. Additional phenotypic assays revealed that Saedanbaek was susceptible to isolate 2457 and that Daewon was susceptible to isolate 2858, indicating that RpsSDB and RpsDW are different genes or alleles that confer race-specific resistance to the two P. sojae isolates. These results provide information that will be helpful for breeders developing P. sojae-resistant cultivars.

Single amino acid change alters specificity of the multi-allelic wheat stem rust resistance locus SR9

Most rust resistance genes thus far isolated from wheat have a very limited number of functional alleles. Here, we report the isolation of most of the alleles at wheat stem rust resistance gene locus SR9. The seven previously reported resistance alleles (Sr9a, Sr9b, Sr9d, Sr9e, Sr9f, Sr9g, and Sr9h) are characterised using a synergistic strategy. Loss-of-function mutants and/or transgenic complementation are used to confirm Sr9b, two haplotypes of Sr9e (Sr9e_h1 and Sr9e_h2), Sr9g, and Sr9h. Each allele encodes a highly related nucleotide-binding site leucine-rich repeat (NB-LRR) type immune receptor, containing an unusual long LRR domain, that confers resistance to a unique spectrum of isolates of the wheat stem rust pathogen. The only SR9 protein effective against stem rust pathogen race TTKSK (Ug99), SR9H, differs from SR9B by a single amino acid. SR9B and SR9G resistance proteins are also distinguished by only a single amino acid. The SR9 allelic series found in the B subgenome are orthologs of wheat stem rust resistance gene Sr21 located in the A subgenome with around 85% identity in protein sequences. Together, our results show that functional diversification of allelic variants at the SR9 locus involves single and multiple amino acid changes that recognize isolates of wheat stem rust.

Comparative genomics profiling revealed multi-stress responsive roles of the CC-NBS-LRR genes in three mango cultivars.

The nucleotide-binding site-leucine-rich repeat (NBS-LRR) gene family is the largest group of disease resistance (R) genes in plants and is active in response to viruses, bacteria, and fungi usually involved in effector-triggered immunity (ETI). Pangenome-wide studies allow researchers to analyze the genetic diversity of multiple species or their members simultaneously, providing a comprehensive understanding of the evolutionary relationships and diversity present among them. The draft pan-genome of three Mangifera indica cultivars (Alphonso, Hong Xiang Ya, and Tommy atkins) was constructed and Presence/absence variants (PAVs) were filtered through the ppsPCP pipeline. As a result, 2823 genes and 5907 PAVs from H. Xiang Ya, and 1266 genes and 2098 PAVs from T. atkins were added to the reference genome. For the identification of CC-NBS-LRR (CNL) genes in these mango cultivars, this draft pan-genome study has successfully identified 47, 27, and 36 members in Alphonso, H. Xiang Ya, and T. atkins respectively. The phylogenetic analysis divided MiCNL proteins into four distinct subgroups. All MiCNL genes are unevenly distributed on chromosomes. Both tandem and segmental duplication events played a significant role in the expansion of the CNL gene family. These genes contain cis-elements related to light, stress, hormone, and development. The analysis of protein-protein interactions (PPI) revealed that MiCNL proteins interacted with other defense-responsive proteins. Gene Ontology (GO) analysis indicated that MiCNL genes play a role in defense mechanisms within the organism. The expression level of the identified genes in fruit peel was observed under disease and cold stress which showed that Mi_A_CNL13 and 14 were up-regulated while Mi_A_CNL15, 25, 30, 31, and 40 were down-regulated in disease stress. On the other hand, Mi_A_CNL2, 14, 41, and 45 were up-regulated and Mi_A_CNL47 is down-regulated in cold stress. Subsequently, the Random Forest (RF) classifier was used to assess the multi-stress response of MiCNLs. It was found that Mi_A_CNL14 is a gene that responds to multiple stress conditions. The CNLs have similar protein structures which show that they are involved in the same function. The above findings provide a foundation for a deeper understanding of the functional characteristics of the mango CNL gene family.

Integrative Molecular Approaches to Plant Disease: A Review

Modern molecular and bioinformatics technologies have made understanding host-pathogen interactions easier. Plants have many ways to protect themselves from microbial diseases, such as physical barriers, PAMP detection, and R genes that recognize pathogen-effector proteins and turn on effector-triggered immunity. Plant pathogen genome databases provide genomic and phenotypic data on plant pathogen species and information on plant-pathogen interactions. Map-based or positional gene cloning is improving our understanding of plant-pathogen interactions, with R genes being used to develop resistance to pathogens. Plant genomes typically contain several hundred nucleotide-binding site-leucine-rich repeats (NLRs), with their number, arrangement, and domain combinations varying by species. Bacterial blight (BB) severely impacts rice production, and about 37 of 44 resistance genes have been mapped and 15 cloned. Many disease-resistant wheat cultivars have been developed using powdery mildew leaf rust (Lr) resistance genes from wild relatives of T. aestivum. Over 140 genes are linked to powdery mildew resistance in T. aestivum and MutChromSeq have found new target genes. Cloning Arabidopsis resistance genes is essential for developing resistant cultivars and understanding R gene evolution. Some R genes encode proteins with nucleotide-binding site (NBS) motifs, and an LRR protects against Erysiphe cruciferarum powdery mildew. CRISPR/Cas9 gene editing is a major tool in plant genome editing, efficiently introducing target site mutations and improving plant immunity. High-throughput sequencing can identify and clone candidate resistance genes in different plant species, and gene editing technologies like CRISPR/Cas have illuminated site-specific mutagenesis and durable resistance.

Deciphering peanut complex genomes paves a way to understand its origin and domestication.

Peanut (Arachis) is a key oil and protein crop worldwide with large genome. The genomes of diploid and tetraploid peanuts have been sequenced, which were compared to decipher their genome structures, evolutionary, and life secrets. Genome sequencing efforts showed that different cultivars, although Bt homeologs being more privileged in gene retention and gene expression. This subgenome bias, extended to sequence variation and point mutation, might be related to the long terminal repeat (LTR) explosions after tetraploidization, especially in At subgenomes. Except that, whole-genome sequences revealed many important genes, for example, fatty acids and triacylglycerols pathway, NBS-LRR (nucleotide-binding site-leucine-rich repeats), and seed size decision genes, were enriched after recursive polyploidization. Each ancestral polyploidy, with old ones having occurred hundreds of thousand years ago, has thousands of duplicated genes in extant genomes, contributing to genetic novelty. Notably, although full genome sequences are available, the actual At subgenome ancestor has still been elusive, highlighted with new debate about peanut origin. Although being an orphan crop lagging behind other crops in genomic resources, the genome sequencing achievement has laid a solid foundation for advancing crop enhancement and system biology research of peanut.

Mapping of the Cladosporium fulvum resistance gene Cf-16, a major gene involved in leaf mold disease in tomato.

Tomato (Solanum lycopersicum) is widely cultivated and consumed worldwide. Tomato leaf mold, caused by Cladosporium fulvum, is one of the most devastating diseases in tomato production. At present, some tomato leaf mold resistance (Cf series) genes used in production gradually lose resistance due to the continuous and rapid differentiation of C. fulvum physiological races. The Cf-16 gene derived from the "Ontario7816" tomato cultivar has shown effective resistance in field trials for many years, but few studies have reported on the mapping of the Cf-16 gene, which has not been cloned, limiting its utilization in tomato breeding. Here, we mapped Cf-16 using a novel comprehensive strategy including bulk segregation analysis (BSA), genome resequencing and SSR molecular markers. A genetic analysis revealed that Cf-16 resistance in "Ontario7816" is controlled by one major dominant locus. The Cf-16 gene was mapped in a region of 2.6cM at chromosome 6 between two markers, namely, TGS447 and TES312, by using an F2 population from a cross between the resistant cultivar "Ontario7816" and susceptible line "Moneymaker." Two nucleotide-binding-site-leucine-rich repeat (NBS-LRR) resistance genes, namely, XM_004240667.3 and XM_010323727.1, were identified in this interval. They are strong candidates for the Cf-16 gene. The mapping of Cf-16 may speed up its utilization for breeding resistant tomato varieties and represents an important step forward in our understanding of the mechanism underlying resistance to tomato leaf mold.

Comparative pangenome analyses provide insights into the evolution of Brassica rapa resistance gene analogues (RGAs).

Brassica rapa is grown worldwide as economically important vegetable and oilseed crop. However, its production is challenged by yield-limiting pathogens. The sustainable control of these pathogens mainly relies on the deployment of genetic resistance primarily driven by resistance gene analogues (RGAs). While several studies have identified RGAs in B. rapa, these were mainly based on a single genome reference and do not represent the full range of RGA diversity in B. rapa. In this study, we utilized the B. rapa pangenome, constructed from 71 lines encompassing 12 morphotypes, to describe a comprehensive repertoire of RGAs in B. rapa. We show that 309 RGAs were affected by presence-absence variation (PAV) and 223 RGAs were missing from the reference genome. The transmembrane leucine-rich repeat (TM-LRR) RGA class had more core gene types than variable genes, while the opposite was observed for nucleotide-binding site leucine-rich repeats (NLRs). Comparative analysis with the B. napus pangenome revealed significant RGA conservation (93%) between the two species. We identified 138 candidate RGAs located within known B. rapa disease resistance QTL, of which the majority were under negative selection. Using blackleg gene homologues, we demonstrated how these genes in B. napus were derived from B. rapa. This further clarifies the genetic relationship of these loci, which may be useful in narrowing-down candidate blackleg resistance genes. This study provides a novel genomic resource towards the identification of candidate genes for breeding disease resistance in B. rapa and its relatives.

PearMODB: a multiomics database forpear (Pyrus) genomics, genetics andbreeding study.

Pear (Pyrus ssp.) belongs to Rosaceae and is an important fruit tree widely cultivated around the world. Currently, challenges to cope with the burgeoning sets of multiomics data are rapidly increasing. Here, we constructed the Pear Multiomics Database (PearMODB) by integrating genome, transcriptome, epigenome and population variation data, and aimed to provide a portal for accessing and analyzing pear multiomics data. A variety of online tools were built including gene search, BLAST, JBrowse, expression heatmap, synteny analysis and primer design. The information of DNA methylation sites and single-nucleotide polymorphisms can be retrieved through the custom JBrowse, providing an opportunity to explore the genetic polymorphisms linked to phenotype variation. Moreover, different gene families involving transcription factors, transcription regulators and disease resistance (nucleotide-binding site leucine-rich repeat) were identified and compiled for quick search. In particular, biosynthetic gene clusters (BGCs) were identified in pear genomes, and specialized webpages were set up to show detailed information of BGCs, laying a foundation for studying metabolic diversity among different pear varieties. Overall, PearMODB provides an important platform for pear genomics, genetics and breeding studies. Database URL http://pearomics.njau.edu.cn.

Identification, characterization, and validation of NBS-encoding genes in grass pea.

Grass pea is a promising crop with the potential to provide food and fodder, but its genomics has not been adequately explored. Identifying genes for desirable traits, such as drought tolerance and disease resistance, is critical for improving the plant. Grass pea currently lacks known R-genes, including the nucleotide-binding site-leucine-rich repeat (NBS-LRR) gene family, which plays a key role in protecting the plant from biotic and abiotic stresses. In our study, we used the recently published grass pea genome and available transcriptomic data to identify 274 NBS-LRR genes. The evolutionary relationships between the classified genes on the reported plants and LsNBS revealed that 124 genes have TNL domains, while 150 genes have CNL domains. All genes contained exons, ranging from 1 to 7. Ten conserved motifs with lengths ranging from 16 to 30 amino acids were identified. We found TIR-domain-containing genes in 132 LsNBSs, with 63 TIR-1 and 69 TIR-2, and RX-CCLike in 84 LsNBSs. We also identified several popular motifs, including P-loop, Uup, kinase-GTPase, ABC, ChvD, CDC6, Rnase_H, Smc, CDC48, and SpoVK. According to the gene enrichment analysis, the identified genes undergo several biological processes such as plant defense, innate immunity, hydrolase activity, and DNA binding. In the upstream regions, 103 transcription factors were identified that govern the transcription of nearby genes affecting the plant excretion of salicylic acid, methyl jasmonate, ethylene, and abscisic acid. According to RNA-Seq expression analysis, 85% of the encoded genes have high expression levels. Nine LsNBS genes were selected for qPCR under salt stress conditions. The majority of the genes showed upregulation at 50 and 200μM NaCl. However, LsNBS-D18, LsNBS-D204, and LsNBS-D180 showed reduced or drastic downregulation compared to their respective expression levels, providing further insights into the potential functions of LsNBSs under salt stress conditions. They provide valuable insights into the potential functions of LsNBSs under salt stress conditions. Our findings also shed light on the evolution and classification of NBS-LRR genes in legumes, highlighting the potential of grass pea. Further research could focus on the functional analysis of these genes, and their potential use in breeding programs to improve the salinity, drought, and disease resistance of this important crop.

Investigation of Resistance Genes in Genus Vigna Reveals Highly Variable NLRome in Parallel Domesticated Member Species.

Vigna is a unique genus that consist of multiple crop species that are domesticated in parallel fashion between 7-10 thousand years ago. Here we studied the evolution of nucleotide-binding site leucine-rich repeat receptor (NLR) genes across five crop species of genus Vigna. In total identified 286, 350, 234, 250, 108 and 161 NLR genes were from Phaseolous vulgaris, Vigna. unguiculata, Vigna mungo, Vigna radiata, Vigna angularis and Vigna umbellata respectively. Comprehensive phylogenetic and clusterization analysis reveals the presence of seven subgroups of Coiled coil like NLRs (CC-NLR) genes and four distinct lineages of Toll interleukin receptor like NLRs (TIR-NLR). Subgroup CCG10-NLR shows large scale diversification among Vigna species suggesting genus specific distinct duplication pattern in Vigna species. Mainly birth of new NLR gene families and higher rate of terminal duplication is the major determinants for expansion of NLRome in genus Vigna. Recent expansion of NLRome in V. anguiculata and V. radiata was also observed which might suggest that domestication have supported their duplication of lineage specific NLR genes. In short, large scale difference in the architecture of NLRome were observed in diploid plant species. Our findings allowed us to hypothesized that independent parallel domestication is the major drivers of highly divergent evolution of NLRome in genus Vigna.