Candidate Genes For Disease Resistance Research Articles

Transcriptome analysis of resistant and susceptible Medicago truncatula genotypes in response to spring black stem and leaf spot disease

Ascochyta blights cause yield losses in all major legume crops. Spring black stem (SBS) and leaf spot disease is a major foliar disease of Medicago truncatula and Medicago sativa (alfalfa) caused by the necrotrophic fungus Ascochyta medicaginicola. This present study sought to identify candidate genes for SBS disease resistance for future functional validation. We employed RNA-seq to profile the transcriptomes of a resistant (HM078) and susceptible (A17) genotype of M. truncatula at 24, 48, and 72 h post inoculation. Preliminary microscopic examination showed reduced pathogen growth on the resistant genotype. In total, 192 and 2,908 differentially expressed genes (DEGs) were observed in the resistant and susceptible genotype, respectively. Functional enrichment analysis revealed the susceptible genotype engaged in processes in the cell periphery and plasma membrane, as well as flavonoid biosynthesis whereas the resistant genotype utilized calcium ion binding, cell wall modifications, and external encapsulating structures. Candidate genes for disease resistance were selected based on the following criteria; among the top ten upregulated or downregulated genes in the resistant genotype, upregulated over time in the resistant genotype, hormone pathway genes, plant disease resistance genes, receptor-like kinases, contrasting expression profiles in QTL for disease resistance, and upregulated genes in enriched pathways. Overall, 22 candidate genes for SBS disease resistance were identified with support from the literature. These genes will be sources for future targeted mutagenesis and candidate gene validation potentially helping to improve disease resistance to this devastating foliar pathogen.

Transcriptomic analysis identifies candidate genes for Aphanomyces root rot disease resistance in pea

BackgroundAphanomyces euteiches is a soil-borne oomycete that causes root rot in pea and other legume species. Symptoms of Aphanomyces root rot (ARR) include root discoloration and wilting, leading to significant yield losses in pea production. Resistance to ARR is known to be polygenic but the roles of single genes in the pea immune response are still poorly understood. This study uses transcriptomics to elucidate the immune response of two pea genotypes varying in their levels of resistance to A. euteiches.ResultsIn this study, we inoculated roots of the pea (P. sativum L.) genotypes ‘Linnea’ (susceptible) and ‘PI180693’ (resistant) with two different A. euteiches strains varying in levels of virulence. The roots were harvested at 6 h post-inoculation (hpi), 20 hpi and 48 hpi, followed by differential gene expression analysis. Our results showed a time- and genotype-dependent immune response towards A. euteiches infection, involving several WRKY and MYB-like transcription factors, along with genes associated with jasmonic acid (JA) and abscisic acid (ABA) signaling. By cross-referencing with genes segregating with partial resistance to ARR, we identified 39 candidate disease resistance genes at the later stage of infection. Among the genes solely upregulated in the resistant genotype ‘PI180693’, Psat7g091800.1 was polymorphic between the pea genotypes and encoded a Leucine-rich repeat receptor-like kinase reminiscent of the Arabidopsis thaliana FLAGELLIN-SENSITIVE 2 receptor.ConclusionsThis study provides new insights into the gene expression dynamics controlling the immune response of resistant and susceptible pea genotypes to A. euteiches infection. We present a set of 39 candidate disease resistance genes for ARR in pea, including the putative immune receptor Psat7g091800.1, for future functional validation.

Genome-wide association study identifies candidate SNPs and genes associated with red-spotted grouper nervous necrosis virus infection of the giant grouper (Epinephelus lanceolatus)

The giant grouper is one of the most important farmed marine animals worldwide; however, red-spotted grouper nervous necrosis virus (RGNNV) has seriously affected its survival. Here, we performed a genome-wide association study on the same pedigree population of 202 giant grouper individuals for full-genome screening to identify markers and candidate genes for RGNNV resistance. The results were used to screen for single-nucleotide polymorphism (SNP) loci and candidate genes that significantly correlated with RGNNV traits. We detected 2,360,340 SNP loci and used a general linear model to identify 54 SNP loci with significant anti-RGNNV associations. We screened for candidate genes in a 100-kb zone (50 kb upstream and 50 kb downstream) around each SNP locus and detected 25 candidate genes. Of these, G protein-coupled receptor 143 (gpr143) was a candidate gene for disease resistance in giant groupers. RT-qPCR was used to examine the expression of gpr143 in various tissues of healthy groupers and was found to be highly expressed in the eye and brain. Following RGNNV infection, gene expression analysis revealed that gpr143 mRNA expression was significantly upregulated in the brains of susceptible fish, suggesting that GPR143 plays a role in viral invasion. GPR143 overexpression in giant grouper cells significantly promoted RGNNV replication, accelerated the cytopathic effects of RGNNV infection, and increased viral gene transcription. Our results suggest that markers linked to RGNNV disease resistance, including SNP markers and genes, could be useful for marker-assisted introgression, allowing for the development of highly RGNNV-resistant giant groupers.

Comparative Transcriptome Analysis Reveals Novel Candidate Resistance Genes Involved in Defence against Phytophthora cactorum in Strawberry.

Crown rot, caused by Phytophthora cactorum, is a devastating disease of strawberry. While most commercial octoploid strawberry cultivars (Fragaria × ananassa Duch) are generally susceptible, the diploid species Fragaria vesca is a potential source of resistance genes to P. cactorum. We previously reported several F. vesca genotypes with varying degrees of resistance to P. cactorum. To gain insights into the strawberry defence mechanisms, comparative transcriptome profiles of two resistant genotypes (NCGR1603 and Bukammen) and a susceptible genotype (NCGR1218) of F. vesca were analysed by RNA-Seq after wounding and subsequent inoculation with P. cactorum. Differential gene expression analysis identified several defence-related genes that are highly expressed in the resistant genotypes relative to the susceptible genotype in response to P. cactorum after wounding. These included putative disease resistance (R) genes encoding receptor-like proteins, receptor-like kinases, nucleotide-binding sites, leucine-rich repeat proteins, RPW8-type disease resistance proteins, and 'pathogenesis-related protein 1'. Seven of these R-genes were expressed only in the resistant genotypes and not in the susceptible genotype, and these appeared to be present only in the genomes of the resistant genotypes, as confirmed by PCR analysis. We previously reported a single major gene locus RPc-1 (Resistance to Phytophthora cactorum 1) in F. vesca that contributed resistance to P. cactorum. Here, we report that 4-5% of the genes (35-38 of ca 800 genes) in the RPc-1 locus are differentially expressed in the resistant genotypes compared to the susceptible genotype after inoculation with P. cactorum. In particular, we identified three defence-related genes encoding wall-associated receptor-like kinase 3, receptor-like protein 12, and non-specific lipid-transfer protein 1-like that were highly expressed in the resistant genotypes compared to the susceptible one. The present study reports several novel candidate disease resistance genes that warrant further investigation for their role in plant defence against P. cactorum.

Genetic diversity and relationships among wild and cultivated Ficus carica L.: Usefulness of RGA markers

Disease resistance and the maintenance of genetic diversity in wild and cultivated populations are very important challenges to implement breeding program and markers assisted selection of Ficus carica L. facing climate change and its consequences. Resistance gene analogs (RGA) markers were used for variety discrimination and assessment of genetic structure and diversity of wild and cultivated Ficus carica L. species in Tunisia. The RGA markers were efficient and reliable markers for discriminating wild and cultivated fig. The high level of polymorphism (95.65) detected suggests the effectiveness of RGAs for both genetic fingerprinting and relationships assessment in wild and cultivated fig. The detected markers may represent candidate genes for disease resistance and could be further used to facilitate the identification of candidate genes and accelerate the genetic improvement of disease resistance in breeding programs of Ficus carica species.

Transcriptomic Analysis Revealed Key Defense Genes and Signaling Pathways Mediated by the Arabidopsis thaliana Gene SAD2 in Response to Infection with Pseudomonas syringae pv. Tomato DC3000.

Nucleocytoplasmic transport receptors play key roles in the nuclear translocation of disease resistance proteins, but the associated mechanisms remain unclear. The Arabidopsis thaliana gene SAD2 encodes an importin β-like protein. A transgenic Arabidopsis line overexpressing SAD2 (OESAD2/Col-0) showed obvious resistance to Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) compared to the wild type (Col-0), but the knockout mutant sad2-5 was susceptible. Transcriptomic analysis was then performed on Col-0, OESAD2/Col-0, and sad2-5 leaves at 0, 1, 2, and 3 days post-inoculation with Pst DC3000. A total of 1825 differentially expressed genes (DEGs) were identified as putative biotic stress defense genes regulated by SAD2, 45 of which overlapped between the SAD2 knockout and overexpression datasets. Gene Ontology (GO) analysis indicated that the DEGs were broadly involved in single-organism cellular metabolic processes and in response to stimulatory stress. Kyoto Encyclopedia of Genes and Genomes (KEGG) biochemical pathway analysis revealed that many of the DEGs were associated with the biosynthesis of flavonoids and other specialized metabolites. Transcription factor analysis showed that a large number of ERF/AP2, MYB, and bHLH transcription factors were involved in SAD2-mediated plant disease resistance. These results provide a basis for future exploration of the molecular mechanisms associated with SAD2-mediated disease resistance and establish a set of key candidate disease resistance genes.

Genome-Wide Analysis of von Willebrand Factor A Gene Family in Rice for Its Role in Imparting Biotic Stress Resistance with Emphasis on Rice Blast Disease

von Willebrand factor A (vWA) genes are well characterized in humans except for few BONZAI genes, but the vWA genes are least explored in plants. Considering the novelty and vital role of vWA genes, this study aimed at characterization of vWA superfamily in rice. Rice genome was found to have 40 vWA genes distributed across all the 12 chromosomes, and 20 of the 40 vWA genes were unique while the remaining shared large fragment similarities with each other, indicating gene duplication. In addition to vWA domain, vWA proteins possess other different motifs or domains, such as ubiquitin interacting motif in protein degradation pathway, and RING finger in protein-protein interaction. Expression analysis of vWA genes in available expression data suggested that they probably function in biotic and abiotic stress responses including hormonal response and signaling. The frequency of transposon elements in the entire 3K rice germplasm was negligible except for 9 vWA genes, indicating the importance of these genes in rice. Structural and functional diversities showed that the vWA genes in a blast-resistant rice variety Tetep had huge variations compared to blast-susceptible rice varieties HP2216 and Nipponbare. qRT-PCR analysis of vWA genes in Magnaporthe oryzae infected rice tissues indicated OsvWA9, OsvWA36, OsvWA37 and OsvWA18 as the optimal candidate genes for disease resistance. This is the first attempt to characterize vWA gene family in plant species.

Construction and characterization of a de novo draft genome of garden cress (Lepidium sativum L.).

Garden cress (Lepidium sativum L.) is a Brassicaceae crop recognized as a healthy vegetable and a medicinal plant. Lepidium is one of the largest genera in Brassicaceae, yet, the genus has not been a focus of extensive genomic research. In the present work, garden cress genome was sequenced using the long read high-fidelity sequencing technology. A de novo, draft genome assembly that spans 336.5Mb was produced, corresponding to 88.6% of the estimated genome size and representing 90% of the evolutionarily expected orthologous gene content. Protein coding gene content was structurally predicted and functionally annotated, resulting in the identification of 25,668 putative genes. A total of 599 candidate disease resistance genes were identified by predicting resistance gene domains in gene structures, and 37 genes were detected as orthologs of heavy metal associated protein coding genes. In addition, 4289 genes were assigned as "transcription factor coding." Six different machine learning algorithms were trained and tested for their performance in classifying miRNA coding genomic sequences. Logistic regression proved the best performing trained algorithm, thus utilized for pre-miRNA coding loci identification in the assembly. Repetitive DNA analysis involved the characterization of transposable element and microsatellite contents. L. sativum chloroplast genome was also assembled and functionally annotated. Data produced in the present work is expected to constitute a foundation for genomic research in garden cress and contribute to genomics-assisted crop improvement and genome evolution studies in the Brassicaceae family.

Quantitative Trait Loci Mapping and Development of KASP Marker Smut Screening Assay Using High-Density Genetic Map and Bulked Segregant RNA Sequencing in Sugarcane (Saccharum spp.).

Sugarcane is one of the most important industrial crops globally. It is the second largest source of bioethanol, and a major crop for biomass-derived electricity and sugar worldwide. Smut, caused by Sporisorium scitamineum, is a major sugarcane disease in many countries, and is managed by smut-resistant varieties. In China, smut remains the single largest constraint for sugarcane production, and consequently it impacts the value of sugarcane as an energy feedstock. Quantitative trait loci (QTLs) associated with smut resistance and linked diagnostic markers are valuable tools for smut resistance breeding. Here, we developed an F1 population (192 progeny) by crossing two sugarcane varieties with contrasting smut resistance and used for genome-wide single nucleotide polymorphism (SNP) discovery and mapping, using a high-throughput genotyping method called “specific locus amplified fragment sequencing (SLAF-seq) and bulked-segregant RNA sequencing (BSR-seq). SLAF-seq generated 148,500 polymorphic SNP markers. Using SNP and previously identified SSR markers, an integrated genetic map with an average 1.96 cM marker interval was produced. With this genetic map and smut resistance scores of the F1 individuals from four crop years, 21 major QTLs were mapped, with a phenotypic variance explanation (PVE) > 8.0%. Among them, 10 QTLs were stable (repeatable) with PVEs ranging from 8.0 to 81.7%. Further, four QTLs were detected based on BSR-seq analysis. aligning major QTLs with the genome of a sugarcane progenitor Saccharum spontaneum, six markers were found co-localized. Markers located in QTLs and functional annotation of BSR-seq-derived unigenes helped identify four disease resistance candidate genes located in major QTLs. 77 SNPs from major QTLs were then converted to Kompetitive Allele-Specific PCR (KASP) markers, of which five were highly significantly linked to smut resistance. The co-localized QTLs, candidate resistance genes, and KASP markers identified in this study provide practically useful tools for marker-assisted sugarcane smut resistance breeding.

A Novel QTL for Resistance to Phytophthora Crown Rot in Squash.

Phytophthora capsici Leonian causes significant yield losses in commercial squash (Cucurbita pepo) production worldwide. The deployment of resistant cultivars can complement integrated management practices for P. capsici, but resistant cultivars are currently unavailable for growers. Moderate resistance to Phytophthora crown rot in a selection of accession PI 181761 (C. pepo) (designated line #181761-36P) is controlled by three dominant genes (R4, R5 and R6). Introgression of these loci into elite germplasm through marker-assisted selection (MAS) can accelerate the release of new C. pepo cultivars resistant to crown rot, but these tools are currently unavailable. Here we describe the identification of a quantitative trait locus (QTL), molecular markers and candidate genes associated with crown rot resistance in #181761-36P. Five hundred and twenty-three SNP markers were genotyped in an F2 (n = 83) population derived from a cross between #181761-36P (R) and Table Queen (S) using targeted genotyping by sequencing. A linkage map (2068.96 cM) consisting of twenty-one linkage groups and an average density of 8.1 markers/cM was developed for the F2 population. The F2:3 families were phenotyped in the greenhouse with a virulent strain of P. capsica, using the spore-spray method. A single QTL (QtlPC-C13) was consistently detected on LG 13 (chromosome 13) across three experiments and explained 17.92–21.47% of phenotypic variation observed in the population. Nine candidate disease resistance gene homologs were found within the confidence interval of QtlPC-C13. Single nucleotide polymorphism (SNP) markers within these genes were converted into Kompetitive Allele Specific PCR (KASP) assays and tested for association with resistance in the F2 population. One SNP marker (C002686) was significantly associated with resistance to crown rot in the F2 population (p < 0.05). This marker is a potential target for MAS for crown rot resistance in C. pepo.

Genome-wide identification and expression patterns in response to signals from &amp;lt;italic&amp;gt;Phytophthora infestans &amp;lt;/italic&amp;gt;of CrRLK1Ls gene family in potato

<p id="C3">Catharanthus roseus receptor-like kinase1-like kinases (CrRLK1Ls), a class of plant-specific protein receptor kinases, play an important role in maintaining cell wall integrity, cell-to-cell communications, and biotic and abiotic stress response. In this study, StCrRLK1Ls family members were identified, and their physical and chemical characteristics, chromosome location, evolutionary characteristics, subcellular location, and the expression patterns were comprehensively analyzed. The results showed that 17 StCrRLK1Ls members were detected, with amino acid size (753-997 aa), molecular weight (83.34-108.69 kD), and isoelectric point (5.30-7.56). Among them, <italic>StCrRLK1Ls</italic> were mainly predicted on plasma membrane. Evolutionary analysis indicated that CrRLK1Ls from potato, rice, apple, <italic>Arabidopsis</italic>, and tomato could be divided into seven subgroups, and <italic>St</italic>CrRLK1Ls were distributed in subgroups III, IV, V, VI, and VII. Moreover, <italic>StCrRLK1Ls </italic>were distributed on eight chromosomes, and there are three tandem repeat gene clusters, including six members. In addition, there were many cis-elements in the promoter region of <italic>StCrRLK1Ls</italic>, which mainly responded to hormones, defense, and stress signals. After the inoculation with <italic>Phytophthora infestans</italic> (<italic>Pi</italic>), a large number of <italic>StCrRLK1Ls</italic> were differentially expressed in Atlantic and Longshu 7, and the relative expression levels of <italic>StCrRLK1L</italic>8 and <italic>StCrRLK1L</italic>10 were obviously up-regulated during <italic>Pi</italic> infection. The relative expression levels of <italic>StCrRLK1L</italic>11 were 9.74 times compared with the control at eight days Atlantic inoculated <italic>Pi</italic>. We speculated that they could be used as the candidate genes for disease resistance and further functional analysis, which might play an important role in response to pathogenic fungi of potato.

High resolution mapping and candidate gene identification of downy mildew race 16 resistance in spinach

BackgroundDowny mildew, the most devastating disease of spinach (Spinacia oleracea L.), is caused by the oomycete Peronospora effusa [=P. farinosa f. sp. spinaciae]. The P. effusa shows race specificities to the resistant host and comprises 19 reported races and many novel isolates. Sixteen new P. effusa races were identified during the past three decades, and the new pathogen races are continually overcoming the genetic resistances used in commercial cultivars. A spinach breeding population derived from the cross between cultivars Whale and Lazio was inoculated with P. effusa race 16 in an environment-controlled facility; disease response was recorded and genotyped using genotyping by sequencing (GBS). The main objective of this study was to identify resistance-associated single nucleotide polymorphism (SNP) markers from the cultivar Whale against the P. effusa race 16.ResultsAssociation analysis conducted using GBS markers identified six significant SNPs (S3_658,306, S3_692697, S3_1050601, S3_1227787, S3_1227802, S3_1231197). The downy mildew resistance locus from cultivar Whale was mapped to a 0.57 Mb region on chromosome 3, including four disease resistance candidate genes (Spo12736, Spo12784, Spo12908, and Spo12821) within 2.69–11.28 Kb of the peak SNP.ConclusionsGenomewide association analysis approach was used to map the P. effusa race 16 resistance loci and identify associated SNP markers and the candidate genes. The results from this study could be valuable in understanding the genetic basis of downy mildew resistance, and the SNP marker will be useful in spinach breeding to select resistant lines.

Frontiers in Dissecting and Managing Brassica Diseases: From Reference-Based RGA Candidate Identification to Building Pan-RGAomes.

The Brassica genus contains abundant economically important vegetable and oilseed crops, which are under threat of diseases caused by fungal, bacterial and viral pathogens. Resistance gene analogues (RGAs) are associated with quantitative and qualitative disease resistance and the identification of candidate RGAs associated with disease resistance is crucial for understanding the mechanism and management of diseases through breeding. The availability of Brassica genome assemblies has greatly facilitated reference-based quantitative trait loci (QTL) mapping for disease resistance. In addition, pangenomes, which characterise both core and variable genes, have been constructed for B. rapa, B. oleracea and B. napus. Genome-wide characterisation of RGAs using conserved domains and motifs in reference genomes and pangenomes reveals their clustered arrangements and presence of structural variations. Here, we comprehensively review RGA identification in important Brassica genome and pangenome assemblies. Comparison of the RGAs in QTL between resistant and susceptible individuals allows for efficient identification of candidate disease resistance genes. However, the reference-based QTL mapping and RGA candidate identification approach is restricted by the under-represented RGA diversity characterised in the limited number of Brassica assemblies. The species-wide repertoire of RGAs make up the pan-resistance gene analogue genome (pan-RGAome). Building a pan-RGAome, through either whole genome resequencing or resistance gene enrichment sequencing, would effectively capture RGA diversity, greatly expanding breeding resources that can be utilised for crop improvement.

“A reference genome assembly and adaptive trait analysis of Castanea mollissima ‘Vanuxem,’ a source of resistance to chestnut blight in restoration breeding”

Forest tree species are increasingly subject to severe mortalities from exotic pests, pathogens, and invasive organisms, accelerated by climate change. Such forest health issues are threatening multiple species and ecosystem sustainability globally. One of the most extreme examples of forest ecosystem disruption is the extirpation of the American chestnut (Castanea dentata) caused by the introduction of chestnut blight and root rot pathogens from Asia. Asian species of chestnut are being employed as donors of disease resistance genes to restore native chestnut species in North America and Europe. To aid in the restoration of threatened chestnut species, we present the assembly of a reference genome for Chinese chestnut (C. mollissima) Vanuxem, one of the donors of disease resistance for American chestnut restoration. From the de novo assembly of the complete genome (725.2 Mb in 14,110 contigs), over half of the sequences have been anchored to the 12 genetic linkage groups. The anchoring is validated by genetic maps and in situ hybridization to chromosomes. We demonstrate the value of the genome as a platform for research and species restoration, including signatures of selection differentiating American chestnut from Chinese chestnut to identify important candidate genes for disease resistance, comparisons of genome organization with other woody species, and a genome-wide examination of progress in backcross breeding for blight resistance. This reference assembly should prove of great value in the understanding, improvement, and restoration of chestnut species.

The WRKY transcription factor PlWRKY65 enhances the resistance of Paeonia lactiflora (herbaceous peony) to Alternaria tenuissima

In this study, the disease resistance gene PlWRKY65 was isolated from the leaves of Paeonia lactiflora and analyzed by bioinformatics methods, and the localization of the encoded protein was explored. Quantitative real-time PCR (qRT-PCR) was also used to explore the response of this gene to Alternaria tenuissima. The results showed that the gene sequence contained multiple cis-acting elements involved in the response to hormone signaling molecules belonging to the IIe subgroup of the WRKY family, and the encoded proteins were located in the nucleus. The PlWRKY65 gene has a positive regulatory effect on A. tenuissima infection. After silencing the PlWRKY65 gene via virus-induced gene silencing (VIGS), it was found that the gene-silenced plants were more sensitive to A. tenuissima infection than the wild plants, exhibiting more severe infection symptoms and different degrees of changes in the expression of the pathogenesis-related (PR) genes. In addition, we showed that the endogenous jasmonic acid (JA) content of P. lactiflora was increased in response to A. tenuissima infection, whereas the salicylic acid (SA) content decreased. After PlWRKY65 gene silencing, the levels of the two hormones changed accordingly, indicating that PlWRKY65, acting as a disease resistance-related transcriptional activator, exerts a regulatory effect on JA and SA signals. This study lays the foundation for functional research on WRKY genes in P. lactiflora and for the discovery of candidate disease resistance genes.

Report on the Thirty-Fifth Southern Forest Tree Improvement Conference (SFTIC 2019)

The 35th biennial Southern Forest Tree Improvement Conference (SFTIC) was held on the 4th and 5th day of June 2019, in Lexington, KY, USA. The theme of the conference was “Genetics and improvement of forest health and productivity”. Plenary speakers detailed significant progress in understanding forest tree genomes, identifying candidate genes for disease resistance and tree architecture traits and utilizing both genomic selection and genetic modification to advance forest health and productivity. Concurrent and general technical sessions provided additional evidence of progress in these areas as well as information on seed zone development for the eastern USA and species prioritization for conservation and restoration programs, in addition to advanced genome-assisted selection and breeding methods and programs in species of commercial interest including pines and eucalyptus. An interactive poster session provided each poster presenter an opportunity to speak in front of a general session audience and to take Q&A in a traditional poster session format. Pre-conference workshops and a post-conference field tour rounded out the 35th SFTIC with 88 registered for the conference, more than 60 participating in workshops and 33 attending the field tour. The conference program and proceedings are accessible through the SFTIC web site at http://www.sftic.org.

Analysis of pooled genome sequences from Djallonke and Sahelian sheep of Ghana reveals co-localisation of regions of reduced heterozygosity with candidate genes for disease resistance and adaptation to a tropical environment

BackgroundThe Djallonke sheep is well adapted to harsh environmental conditions, and is relatively resistant to Haemonchosis and resilient to animal trypanosomiasis. The larger Sahelian sheep, which cohabit the same region, is less well adapted to these disease challenges. Haemonchosis and Trypanosomiasis collectively cost the worldwide animal industry billions of dollars in production losses annually.ResultsHere, we separately sequenced and then pooled according to breed the genomes from five unrelated individuals from each of the Djallonke and Sahelian sheep breeds (sourced from Ghana), at greater than 22-fold combined coverage for each breed. A total of approximately 404 million (97%) and 343 million (97%) sequence reads from the Djallonke and Sahelian breeds respectively, were successfully mapped to the sheep reference genome Oar v3.1. We identified approximately 11.1 million and 10.9 million single nucleotide polymorphisms (SNPs) in the Djallonke and Sahelian breeds, with approximately 15 and 16% respectively of these not previously reported in sheep. Multiple regions of reduced heterozygosity were also found; 70 co-localised within genomic regions harbouring genes that mediate disease resistance, immune response and adaptation in sheep or cattle. Thirty- three of the regions of reduced heterozygosity co-localised with previously reported genes for resistance to haemonchosis and trypanosomiasis.ConclusionsOur analyses suggest that these regions of reduced heterozygosity may be signatures of selection for these economically important diseases.

Sequence characterization, structural analysis, SNP detection and expression profiling of SLC11A1 gene in Indian goats

The Solute Carrier Family 11 Member A1 (SLC11A1) is a candidate gene, encodes a divalent cation transporter, SLC11A1 protein, localizes in the phagolysosome membrane in macrophages, essential for immunity and resistance to intracellular pathogens. The indigenous goats of Kerala are known for disease resistance and adaptability, but the underlying genetic causes are to be unraveled. This study comprised of characterization of the caprine SLC11A1 gene, to detect SNPs and compare the relative abundance of SLC11A1 mRNA by quantitative-PCR in native goats (Attappady Black and Malabari) and crossbreds. The 1702 bp cDNA of SLC11A1 comprised of 15 exons, an ORF of 1647 bp, encoding a protein of 548 amino acids. SLC11A1 amino acid sequence was 83–99% identical to that of other species. SLC11A1 peptide contains 10 serine and 2 threonine phosphorylation loci and 12 transmembrane domains. The study indicated the presence of four synonymous and three non-synonymous mutations (p.Ile132Thr; p.Phe294Leu; p.Gln541His). The amino acid substitution p.Ile132Thr was deleterious and p.Phe294Leu and p.Gln541His were neutral. p.Phe294Leu was located in the 7th transmembrane helix. The relative abundance of the SLC11A1 mRNA in leukocytes in Malabari and Attappady Black were significantly higher than crossbreds (P < 0.01). The significantly higher SLC11A1 expression in native goats indicates higher disease resistance than crossbreds, gives an insight into the necessity of conservation of native breeds, important role of SLC11A1 in disease resistance. The detected SNPs would benefit the use of this gene as a candidate gene for disease resistance in goat breeding programs.

Genetic mapping and haplotype analysis of a locus for quantitative resistance to Fusarium graminearum in soybean accession PI 567516C.

A major novel quantitative disease resistance locus, qRfg_Gm06, for Fusarium graminearum was genetically mapped to chromosome 6. Genomic-assisted haplotype analysis within this region identified three putative candidate genes. Fusarium graminearum causes seed, root rot, and seedling damping-off in soybean which contributes to reduced stands and yield. A cultivar Magellan and PI 567516C were identified with low and high levels of partial resistance to F. graminearum, respectively. Quantitative disease resistance loci (QDRL) were mapped with 241 F7:8 recombinant inbred lines (RILs) derived from a cross of Magellan × PI 567516C. Phenotypic evaluation for resistance to F. graminearum used the rolled towel assay in a randomized incomplete block design. The genetic map was constructed from 927 polymorphic single nucleotide polymorphism (SNP) and simple sequence repeat (SSR) markers. One major QDRL qRfg_Gm06 was detected and mapped to chromosome 6 with a LOD score of 20.3 explaining 40.2% of the total phenotypic variation. This QDRL was mapped to a ~400kb genomic region of the Williams 82 reference genome. Genome mining of this region identified 14 putative candidate disease resistance genes. Haplotype analysis of this locus using whole genome re-sequencing (WGRS) of 106 diverse soybean lines narrowed the list to three genes. A SNP genotyping Kompetitive allele-specific PCR (KASP) assay was designed for one of the genes and was validated in a subset of the RILs and all 106 diverse lines.

Fine mapping of the stem rust resistance gene SrTA10187.

SrTA10187 was fine-mapped to a 1.1cM interval, candidate genes were identified in the region of interest, and molecular markers were developed for marker-assisted selection and Sr gene pyramiding. Stem rust (Puccinia graminis f. sp. tritici, Pgt) races belonging to the Ug99 (TTKSK) race group pose a serious threat to global wheat (Triticum aestivum L.) production. To improve Pgt host resistance, the Ug99-effective resistance gene SrTA10187 previously identified in Aegilops tauschii Coss. was introgressed into wheat, and mapped to the short arm of wheat chromosome 6D. In this study, high-resolution mapping of SrTA10187 was done using a population of 1,060 plants. Pgt resistance was screened using race QFCSC. PCR-based SNP and STS markers were developed from genotyping-by-sequencing tags and SNP sequences available in online databases. SrTA10187 segregated as expected in a 3:1 ratio of resistant to susceptible individuals in three out of six BC3F2 families, and was fine-mapped to a 1.1cM region on wheat chromosome 6DS. Marker context sequence was aligned to the reference Ae. tauschii genome to identify the physical region encompassing SrTA10187. Due to the size of the corresponding region, candidate disease resistance genes could not be identified with confidence. Comparisons with the Ae. tauschii genetic map developed by Luo et al. (PNAS 110(19):7940-7945, 2013) enabled identification of a discrete genetic locus and a BAC minimum tiling path of the region spanning SrTA10187. Annotation of pooled BAC library sequences led to the identification of candidate genes in the region of interest-including a single NB-ARC-LRR gene. The shorter genetic interval and flanking KASP™ and STS markers developed in this study will facilitate marker-assisted selection, gene pyramiding, and positional cloning of SrTA10187.