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Horizon scans have emerged as a valuable tool to anticipate the incoming invasive alien species (IAS) by judging species on their potential impacts. However, little research has been conducted on quantifying actual impacts and assessing causes of species-specific vulnerabilities to particular IAS due to persistent methodological challenges. The underlying interspecific mechanisms driving species-specific vulnerabilities therefore remain poorly understood, even though they can substantially improve the accuracy of risk assessments. Given that interspecific interactions underlying ecological impacts of IAS are often shaped by phenological synchrony, we tested the hypothesis that temporal mismatches in breeding phenology between native species and IAS can mitigate their ecological impacts. Focusing on the invasive American bullfrog (Lithobates catesbeianus), we combined an environmental DNA (eDNA) quantitative barcoding and metabarcoding survey in Belgium with a global meta-analysis, and integrated citizen-science data on breeding phenology. We examined whether the presence of native amphibian species was negatively related to the presence or abundance of invasive bullfrogs and whether this relationship was affected by their phenological mismatches. The field study revealed a significant negative effect of increasing bullfrog eDNA concentrations on native amphibian species richness and community structure. These observations were shaped by species-specific vulnerabilities to invasive bullfrogs, with late spring- and summer-breeding species being strongly affected, while winter-breeding species remained unaffected. This trend was confirmed by the global meta-analysis. A significant negative relationship was observed between phenological mismatch and the impact of bullfrogs. Specifically, native amphibian species with breeding phenology differing by 6 weeks or less from invasive bullfrogs were more likely to be absent in the presence of bullfrogs than species whose phenology differed by more than 6 weeks with that of bullfrogs. Taken together, we present a novel method based on the combination of aqueous eDNA quantitative barcoding and metabarcoding to quantify the ecological impacts of biological invaders at the community level. We show that phenological mismatches between native and invasive species can be a strong predictor of invasion impact regardless of ecological or methodological context. Therefore, we advocate for the integration of temporal alignment between native and IAS's phenologies into invasion impact frameworks.
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We genotyped a population of 618 diploid potato clones derived from six independent potato-breeding programmes from NW-Europe. The diploids were phenotyped for 23 traits, using standardised protocols and common check varieties, enabling us to derive whole population estimators for most traits. We subsequently performed a Genome-Wide Association Study (GWAS) to identify quantitative trait loci (QTL) for all traits with SNPs and short-read haplotypes derived from read-backed phasing. In this study, we used a marker platform called PotatoMASH (Potato Multi-Allele Scanning Haplotags); a pooled multiplex amplicon sequencing based approach. Through this method, neighbouring SNPs within an amplicon can be combined to generate multi-allelic short-read haplotypes (haplotags) that capture recombination history between the constituent SNPs, and reflect the allelic diversity of a given locus in a different way than single bi-allelic SNPs. We found a total of 37 unique QTL across both marker types. A core of 10 QTL were detected with SNPs as well as with haplotags. Haplotags allowed to detect an additional 14 QTL not found based on the SNP set. Conversely, the bi-allelic SNP set also found 13 QTL not detectable using the haplotag set. We conclude that both marker types should routinely be used in parallel to maximize the QTL detection power. We report 19 novel QTL for nine traits: Skin Smoothness, Sprout Dormancy, Total Tuber Number, Tuber Length, Yield, Chipping Colour, After-cooking Blackening, Cooking Type and Eye depth.
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Genomic prediction has mostly been used in single environment contexts, largely ignoring genotype x environment interaction, which greatly affects the performance of plants. However, in the last decade, prediction models including marker x environment (MxE) interaction have been developed. We evaluated the potential of genomic prediction in red clover (Trifolium pratense L.) using field trial data from five European locations, obtained in the Horizon 2020 EUCLEG project. Three models were compared: (1) single environment (SingleEnv), (2) across environment (AcrossEnv), (3) marker x environment interaction (MxE). Annual dry matter yield (DMY) gave the highest predictive ability (PA). Joint analyses of DMY from years 1 and 2 from each location varied from 0.87 in Britain and Switzerland in year 1, to 0.40 in Serbia in year 2. Overall, crude protein (CP) was predicted poorly. PAs for date of flowering (DOF), however ranged from 0.87 to 0.67 for Britain and Switzerland, respectively. Across the three traits, the MxE model performed best and the AcrossEnv worst, demonstrating that including marker x environment effects can improve genomic prediction in red clover. Leaving out accessions from specific regions or from specific breeders' material in the cross validation tended to reduce PA, but the magnitude of reduction depended on trait, region and breeders' material, indicating that population structure contributed to the high PAs observed for DMY and DOF. Testing the genomic estimated breeding values on new phenotypic data from Sweden showed that DMY training data from Britain gave high PAs in both years (0.43-0.76), while DMY training data from Switzerland gave high PAs only for year 1 (0.70-0.87). The genomic predictions we report here underline the potential benefits of incorporating MxE interaction in multi-environment trials and could have perspectives for identifying markers with effects that are stable across environments, and markers with environment-specific effects.
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Though facing significant challenges, coffee (Coffea arabica) grown in Haitian agroforestry systems are important contributors to rural livelihoods and provide several ecosystem services. However, little is known about their genetic diversity and the variety mixtures used. In light of this, there is a need to characterize Haitian coffee diversity to help inform revitalization of this sector. We sampled 28 diverse farms in historically important coffee growing regions of northern and southern Haiti. We performed KASP-genotyping of SNP markers and HiPlex multiplex amplicon sequencing for haplotype calling on our samples, as well as several Ethiopian and commercial accessions from international collections. This allowed us to assign Haitian samples to varietal groups. Our analyses revealed considerable genetic diversity in Haitian farms, higher in fact than many farmers realized. Notably, genetic structure analyses revealed the presence of clusters related to Typica, Bourbon, and Catimor groups, another group that was not represented in our reference accession panel, and several admixed individuals. Across the study areas, we found both mixed-variety farms and monovarietal farms with the historical and traditional Typica variety. This study is, to our knowledge, the first to genetically characterize Haitian C. arabica variety mixtures, and report the limited cultivation of C. canephora (Robusta coffee) in the study area. Our results show that some coffee farms are repositories of historical, widely-abandoned varieties while others are generators of new diversity through genetic mixing.
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Coffea , Café , Humanos , Haití , Ecosistema , Coffea/genética , Variación GenéticaRESUMEN
BACKGROUND AND AIMS: Plant breeders are increasingly turning to crop wild relatives (CWRs) to ensure food security in a rapidly changing environment. However, CWR populations are confronted with various human-induced threats, including hybridization with their nearby cultivated crops. This might be a particular problem for wild coffee species, which often occur near coffee cultivation areas. Here, we briefly review the evidence for wild Coffea arabica (cultivated as Arabica coffee) and Coffea canephora (cultivated as Robusta coffee) and then focused on C. canephora in the Yangambi region in the Democratic Republic of the Congo. There, we examined the geographical distribution of cultivated C. canephora and the incidence of hybridization between cultivated and wild individuals within the rainforest. METHODS: We collected 71 C. canephora individuals from home gardens and 12 C. canephora individuals from the tropical rainforest in the Yangambi region and genotyped them using genotyping-by-sequencing (GBS). We compared the fingerprints with existing GBS data from 388 C. canephora individuals from natural tropical rainforests and the INERA Coffee Collection, a Robusta coffee field gene bank and the most probable source of cultivated genotypes in the area. We then established robust diagnostic fingerprints that genetically differentiate cultivated from wild coffee, identified cultivated-wild hybrids and mapped their geographical position in the rainforest. KEY RESULTS: We identified cultivated genotypes and cultivated-wild hybrids in zones with clear anthropogenic activity, and where cultivated C. canephora in home gardens may serve as a source for crop-to-wild gene flow. We found relatively few hybrids and backcrosses in the rainforests. CONCLUSIONS: The cultivation of C. canephora in close proximity to its wild gene pool has led to cultivated genotypes and cultivated-wild hybrids appearing within the natural habitats of C. canephora. Yet, given the high genetic similarity between the cultivated and wild gene pool, together with the relatively low incidence of hybridization, our results indicate that the overall impact in terms of risk of introgression remains limited so far.
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Coffea , Flujo Génico , Coffea/genética , República Democrática del Congo , Productos Agrícolas/genética , Hibridación Genética , Bosque Lluvioso , GenotipoRESUMEN
Marine sediments cover 70% of the Earth's surface, and harbour diverse bacterial communities critical for marine biogeochemical processes, which affect climate change, biodiversity and ecosystem functioning. Nematodes, the most abundant and species-rich metazoan organisms in marine sediments, in turn, affect benthic bacterial communities and bacterial-mediated ecological processes, but the underlying mechanisms by which they affect biogeochemical cycles remain poorly understood. Here, we demonstrate using a metatranscriptomic approach that nematodes alter the taxonomic and functional profiles of benthic bacterial communities. We found particularly strong stimulation of nitrogen-fixing and methane-oxidizing bacteria in the presence of nematodes, as well as increased functional activity associated with methane metabolism and degradation of various carbon compounds. This study provides empirical evidence that the presence of nematodes results in taxonomic and functional shifts in active bacterial communities, indicating that nematodes may play an important role in benthic ecosystem processes.
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Bacterias , Ecosistema , Sedimentos Geológicos , Nematodos , Animales , Nematodos/microbiología , Nematodos/genética , Bacterias/genética , Bacterias/clasificación , Sedimentos Geológicos/microbiología , Biodiversidad , Transcriptoma , Microbiota/genética , Metano/metabolismoRESUMEN
Industrial chicory (Cichorium intybus var. sativum) and witloof (C. intybus var. foliosum) are crops with an important economic value, mainly cultivated for inulin production and as a leafy vegetable, respectively. Both crops are rich in nutritionally relevant specialized metabolites with beneficial effects for human health. However, their bitter taste, caused by the sesquiterpene lactones (SLs) produced in leaves and taproot, limits wider applications in the food industry. Changing the bitterness would thus create new opportunities with a great economic impact. Known genes encoding enzymes involved in the SL biosynthetic pathway are GERMACRENE A SYNTHASE (GAS), GERMACRENE A OXIDASE (GAO), COSTUNOLIDE SYNTHASE (COS) and KAUNIOLIDE SYNTHASE (KLS). In this study, we integrated genome and transcriptome mining to further unravel SL biosynthesis. We found that C. intybus SL biosynthesis is controlled by the phytohormone methyl jasmonate (MeJA). Gene family annotation and MeJA inducibility enabled the pinpointing of candidate genes related with the SL biosynthetic pathway. We specifically focused on members of subclade CYP71 of the cytochrome P450 family. We verified the biochemical activity of 14 C. intybus CYP71 enzymes transiently produced in Nicotiana benthamiana and identified several functional paralogs for each of the GAO, COS and KLS genes, pointing to redundancy in and robustness of the SL biosynthetic pathway. Gene functionality was further analyzed using CRISPR/Cas9 genome editing in C. intybus. Metabolite profiling of mutant C. intybus lines demonstrated a successful reduction in SL metabolite production. Together, this study increases our insights into the C. intybus SL biosynthetic pathway and paves the way for the engineering of C. intybus bitterness.
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China's and Europe's dependence on imported protein is a threat to the food self-sufficiency of these regions. It could be solved by growing more legumes, including alfalfa that is the highest protein producer under temperate climate. To create productive and high-value varieties, the use of large genetic diversity combined with genomic evaluation could improve current breeding programs. To study alfalfa diversity, we have used a set of 395 alfalfa accessions (i.e. populations), mainly from Europe, North and South America and China, with fall dormancy ranging from 3 to 7 on a scale of 11. Five breeders provided materials (617 accessions) that were compared to the 400 accessions. All accessions were genotyped using Genotyping-by-Sequencing (GBS) to obtain SNP allele frequency. These genomic data were used to describe genetic diversity and identify genetic groups. The accessions were phenotyped for phenology traits (fall dormancy and flowering date) at two locations (Lusignan in France, Novi Sad in Serbia) from 2018 to 2021. The QTL were detected by a Multi-Locus Mixed Model (mlmm). Subsequently, the quality of the genomic prediction for each trait was assessed. Cross-validation was used to assess the quality of prediction by testing GBLUP, Bayesian Ridge Regression (BRR), and Bayesian Lasso methods. A genetic structure with seven groups was found. Most of these groups were related to the geographical origin of the accessions and showed that European and American material is genetically distinct from Chinese material. Several QTL associated with fall dormancy were found and most of these were linked to genes. In our study, the infinitesimal methods showed a higher prediction quality than the Bayesian Lasso, and the genomic prediction achieved high (>0.75) predicting abilities in some cases. Our results are encouraging for alfalfa breeding by showing that it is possible to achieve high genomic prediction quality.
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Industrial chicory (Cichorium intybus var. sativum) is a biannual crop mostly cultivated for extraction of inulin, a fructose polymer used as a dietary fiber. F1 hybrid breeding is a promising breeding strategy in chicory but relies on stable male sterile lines to prevent self-pollination. Here, we report the assembly and annotation of a new industrial chicory reference genome. Additionally, we performed RNA-Seq on subsequent stages of flower bud development of a fertile line and two cytoplasmic male sterile (CMS) clones. Comparison of fertile and CMS flower bud transcriptomes combined with morphological microscopic analysis of anthers, provided a molecular understanding of anther development and identified key genes in a range of underlying processes, including tapetum development, sink establishment, pollen wall development and anther dehiscence. We also described the role of phytohormones in the regulation of these processes under normal fertile flower bud development. In parallel, we evaluated which processes are disturbed in CMS clones and could contribute to the male sterile phenotype. Taken together, this study provides a state-of-the-art industrial chicory reference genome, an annotated and curated candidate gene set related to anther development and male sterility as well as a detailed molecular timetable of flower bud development in fertile and CMS lines.
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A major advantage of using CRISPR/Cas9 for gene editing is multiplexing, that is, the simultaneous targeting of many genes. However, primary transformants typically contain hetero-allelic mutations or are genetic mosaic, while genetically stable lines that are homozygous are desired for functional analysis. Currently, a dedicated and labor-intensive effort is required to obtain such higher-order mutants through several generations of genetic crosses and genotyping. We describe the design and validation of a rapid and efficient strategy to produce lines of genetically identical plants carrying various combinations of homozygous edits, suitable for replicated analysis of phenotypical differences. This approach was achieved by combining highly multiplex gene editing in Zea mays (maize) with in vivo haploid induction and efficient in vitro generation of doubled haploid plants using embryo rescue doubling. By combining three CRISPR/Cas9 constructs that target in total 36 genes potentially involved in leaf growth, we generated an array of homozygous lines with various combinations of edits within three generations. Several genotypes show a reproducible 10% increase in leaf size, including a septuple mutant combination. We anticipate that our strategy will facilitate the study of gene families via multiplex CRISPR mutagenesis and the identification of allele combinations to improve quantitative crop traits.
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Edición Génica , Zea mays , Zea mays/genética , Sistemas CRISPR-Cas/genética , Genoma de Planta , Haploidia , Plantas Modificadas GenéticamenteRESUMEN
Improvement of persistency is an important breeding goal in red clover (Trifolium pratense L.). In areas with cold winters, lack of persistency is often due to poor winter survival, of which low freezing tolerance (FT) is an important component. We conducted a genome wide association study (GWAS) to identify loci associated with freezing tolerance in a collection of 393 red clover accessions, mostly of European origin, and performed analyses of linkage disequilibrium and inbreeding. Accessions were genotyped as pools of individuals using genotyping-by-sequencing (pool-GBS), generating both single nucleotide polymorphism (SNP) and haplotype allele frequency data at accession level. Linkage disequilibrium was determined as a squared partial correlation between the allele frequencies of pairs of SNPs and found to decay at extremely short distances (< 1 kb). The level of inbreeding, inferred from the diagonal elements of a genomic relationship matrix, varied considerably between different groups of accessions, with the strongest inbreeding found among ecotypes from Iberia and Great Britain, and the least found among landraces. Considerable variation in FT was found, with LT50-values (temperature at which 50% of the plants are killed) ranging from -6.0°C to -11.5°C. SNP and haplotype-based GWAS identified eight and six loci significantly associated with FT (of which only one was shared), explaining 30% and 26% of the phenotypic variation, respectively. Ten of the loci were found within or at a short distance (<0.5 kb) from genes possibly involved in mechanisms affecting FT. These include a caffeoyl shikimate esterase, an inositol transporter, and other genes involved in signaling, transport, lignin synthesis and amino acid or carbohydrate metabolism. This study paves the way for a better understanding of the genetic control of FT and for the development of molecular tools for the improvement of this trait in red clover through genomics assisted breeding.
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Monitoring fish communities is central to the evaluation of ecological health of rivers. Both presence/absence of fish species and their relative quantity in local fish assemblages are crucial parameters to measure. Fish communities in lotic systems are traditionally monitored via electrofishing, characterized by a known limited efficiency and high survey costs. Analysis of environmental DNA could serve as a non-destructive alternative for detection and quantification of lotic fish communities, but this approach still requires further insights in practical sampling schemes incorporating transport and dilution of the eDNA particles; optimization of predictive power and quality assurance of the molecular detection method. Via a controlled cage experiment, we aim to extend the knowledge on streamreach of eDNA in small rivers and large brooks, as laid out in the European Water Framework Directive's water typology. Using a high and low source biomass in two river transects of a species-poor river characterized by contrasting river discharge rates, we found strong and significant correlations between the eDNA relative species abundances and the relative biomass per species in the cage community. Despite a decreasing correlation over distance, the underlying community composition remained stable from 25 to 300 m, or up to 1 km downstream of the eDNA source, depending on the river discharge rate. Such decrease in similarity between relative source biomass and the corresponding eDNA-based community profile with increasing distance downstream from the source, might be attributed to variation in species-specific eDNA persistence. Our findings offer crucial insights on eDNA behaviour and characterization of riverine fish communities. We conclude that water sampled from a relatively small river offers an adequate eDNA snapshot of the total fish community in the 300-1000 m upstream transect. The potential application for other river systems is further discussed.
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ADN Ambiental , Animales , Biodiversidad , Código de Barras del ADN Taxonómico/métodos , Monitoreo del Ambiente/métodos , Peces/genética , Agua , EcosistemaRESUMEN
Red clover (Trifolium pratense L.) is an outcrossing forage legume that has adapted to a wide range of climatic and growing conditions across Europe. Red clover is valued for its high yield potential and its forage quality. The high amount of genetic diversity present in red clover provides an invaluable, but often poorly characterized resource to improve key traits such as yield, quality, and resistance to biotic and abiotic stresses. In this study, we examined the genetic and phenotypic diversity within a diverse set of 395 diploid red clover accessions via genome wide allele frequency fingerprinting and multi-location field trials across Europe. We found that the genetic structure of accessions mostly reflected their geographic origin and only few cases were detected, where breeders integrated foreign genetic resources into their local breeding pools. The mean dry matter yield of the first main harvesting season ranged from 0.74 kg m-2 in Serbia and Norway to 1.34 kg m-2 in Switzerland. Phenotypic performance of accessions in the multi-location field trials revealed a very strong accession x location interaction. Local adaptation was especially prominent in Nordic red clover accessions that showed a distinct adaptation to the growing conditions and cutting regime of the North. The traits vigor, dry matter yield and plant density were negatively correlated between the trial location in Norway and the locations Great Britain, Switzerland, Czech Republic and Serbia. Notably, breeding material and cultivars generally performed well at the location where they were developed. Our results confirmed that red clover cultivars were bred from regional ecotypes and show a narrow adaptation to regional conditions. Our study can serve as a valuable basis for identifying interesting materials that express the desired characteristics and contribute to the adaptation of red clover to future climatic conditions.
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Degradation and regeneration of tropical forests can strongly affect gene flow in understorey species, resulting in genetic erosion and changes in genetic structure. Yet, these processes remain poorly studied in tropical Africa. Coffea canephora is an economically important species, found in the understorey of tropical rainforests of Central and West Africa, and the genetic diversity harboured in its wild populations is vital for sustainable coffee production worldwide. Here, we aimed to quantify genetic diversity, genetic structure, and pedigree relations in wild C. canephora populations, and we investigated associations between these descriptors and forest disturbance and regeneration. Therefore, we sampled 256 C. canephora individuals within 24 plots across three forest categories in Yangambi (DR Congo), and used genotyping-by-sequencing to identify 18,894 SNPs. Overall, we found high genetic diversity, and no evidence of genetic erosion in C. canephora in disturbed old-growth forest, as compared to undisturbed old-growth forest. In addition, an overall heterozygosity excess was found in all populations, which was expected for a self-incompatible species. Genetic structure was mainly a result of isolation-by-distance, reflecting geographical location, with low to moderate relatedness at finer scales. Populations in regrowth forest had lower allelic richness than populations in old-growth forest and were characterised by a lower inter-individual relatedness and a lack of isolation-by-distance, suggesting that they originated from different neighbouring populations and were subject to founder effects. Wild Robusta coffee populations in the study area still harbour high levels of genetic diversity, yet careful monitoring of their response to ongoing forest degradation remains required.
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Coffea , Humanos , Coffea/genética , Café , República Democrática del Congo , Bosques , Variación GenéticaRESUMEN
Multiplex amplicon sequencing is a versatile method to identify genetic variation in natural or mutagenized populations through eco-tilling or multiplex CRISPR screens. Such genotyping screens require reliable and specific primer designs, combined with simultaneous gRNA design for CRISPR screens. Unfortunately, current tools are unable to combine multiplex gRNA and primer design in a high-throughput and easy-to-use manner with high design flexibility. Here, we report the development of a bioinformatics tool called SMAP design to overcome these limitations. We tested SMAP design on several plant and non-plant genomes and obtained designs for more than 80-90% of the target genes, depending on the genome and gene family. We validated the designs with Illumina multiplex amplicon sequencing and Sanger sequencing in Arabidopsis, soybean, and maize. We also used SMAP design to perform eco-tilling by tilling PCR amplicons across nine candidate genes putatively associated with haploid induction in Cichorium intybus. We screened 60 accessions of chicory and witloof and identified thirteen knockout haplotypes and their carriers. SMAP design is an easy-to-use command-line tool that generates highly specific gRNA and/or primer designs for any number of loci for CRISPR or natural variation screens and is compatible with other SMAP modules for seamless downstream analysis.
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Variación Genética , Reacción en Cadena de la Polimerasa Multiplex , Programas Informáticos , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Sistemas CRISPR-Cas , Genoma , GenotipoRESUMEN
Conventional wisdom states that genetic variation reduces disease levels in plant populations. Nevertheless, crop species have been subject to a gradual loss of genetic variation through selection for specific traits during breeding, thereby increasing their vulnerability to biotic stresses such as pathogens. We explored how genetic variation in Arabica coffee sites in southwestern Ethiopia was related to the incidence of four major fungal diseases. Sixty sites were selected along a gradient of management intensity, ranging from nearly wild to intensively managed coffee stands. We used genotyping-by-sequencing of pooled leaf samples (pool-GBS) derived from 16 individual coffee shrubs in each of the 60 sites to assess the variation in genetic composition (multivariate: reference allele frequency) and genetic diversity (univariate: mean expected heterozygosity) between sites. We found that genetic composition had a clear spatial pattern and that genetic diversity was higher in less managed sites. The incidence of the four fungal diseases was related to the genetic composition of the coffee stands, but in a specific way for each disease. In contrast, genetic diversity was only related to the within-site variation of coffee berry disease, but not to the mean incidence of any of the four diseases across sites. Given that fungal diseases are major challenges of Arabica coffee in its native range, our findings that genetic composition of coffee sites impacted the major fungal diseases may serve as baseline information to study the molecular basis of disease resistance in coffee. Overall, our study illustrates the need to consider both host genetic composition and genetic diversity when investigating the genetic basis for variation in disease levels.
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Coffea , Micosis , Coffea/genética , Fitomejoramiento , EtiopíaRESUMEN
Ensuring food security for an ever-growing global population while adapting to climate change is the main challenge for agriculture in the 21st century. Although new technologies are being applied to tackle this problem, we are approaching a plateau in crop improvement using conventional breeding. Recent advances in CRISPR/Cas9-mediated gene engineering have paved the way to accelerate plant breeding to meet this increasing demand. However, many traits are governed by multiple small-effect genes operating in complex interactive networks. Here, we present the gene discovery pipeline BREEDIT, which combines multiplex genome editing of whole gene families with crossing schemes to improve complex traits such as yield and drought tolerance. We induced gene knockouts in 48 growth-related genes into maize (Zea mays) using CRISPR/Cas9 and generated a collection of over 1,000 gene-edited plants. The edited populations displayed (on average) 5%-10% increases in leaf length and up to 20% increases in leaf width compared with the controls. For each gene family, edits in subsets of genes could be associated with enhanced traits, allowing us to reduce the gene space to be considered for trait improvement. BREEDIT could be rapidly applied to generate a diverse collection of mutants to identify promising gene modifications for later use in breeding programs.
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Edición Génica , Zea mays , Zea mays/genética , Sistemas CRISPR-Cas/genética , Plantas Modificadas Genéticamente/genética , Herencia Multifactorial , Fitomejoramiento , Genoma de Planta/genéticaRESUMEN
KEY MESSAGE: High variability for and candidate loci associated with resistance to southern anthracnose and clover rot in a worldwide collection of red clover provide a first basis for genomics-assisted breeding. Red clover (Trifolium pratense L.) is an important forage legume of temperate regions, particularly valued for its high yield potential and its high forage quality. Despite substantial breeding progress during the last decades, continuous improvement of cultivars is crucial to ensure yield stability in view of newly emerging diseases or changing climatic conditions. The high amount of genetic diversity present in red clover ecotypes, landraces, and cultivars provides an invaluable, but often unexploited resource for the improvement of key traits such as yield, quality, and resistance to biotic and abiotic stresses. A collection of 397 red clover accessions was genotyped using a pooled genotyping-by-sequencing approach with 200 plants per accession. Resistance to the two most pertinent diseases in red clover production, southern anthracnose caused by Colletotrichum trifolii, and clover rot caused by Sclerotinia trifoliorum, was assessed using spray inoculation. The mean survival rate for southern anthracnose was 22.9% and the mean resistance index for clover rot was 34.0%. Genome-wide association analysis revealed several loci significantly associated with resistance to southern anthracnose and clover rot. Most of these loci are in coding regions. One quantitative trait locus (QTL) on chromosome 1 explained 16.8% of the variation in resistance to southern anthracnose. For clover rot resistance we found eight QTL, explaining together 80.2% of the total phenotypic variation. The SNPs associated with these QTL provide a promising resource for marker-assisted selection in existing breeding programs, facilitating the development of novel cultivars with increased resistance against two devastating fungal diseases of red clover.
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Sitios de Carácter Cuantitativo , Trifolium , Trifolium/genética , Medicago/genética , Estudio de Asociación del Genoma Completo , Fitomejoramiento , Variación Biológica Poblacional , Resistencia a la Enfermedad/genética , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiologíaRESUMEN
MAIN CONCLUSION: Through selective genotyping of pooled phenotypic extremes, we identified a number of loci and candidate genes putatively controlling timing of stem elongation in red clover. We have identified candidate genes controlling the timing of stem elongation prior to flowering in red clover (Trifolium pratense L.). This trait is of ecological and agronomic significance, as it affects fitness, competitivity, climate adaptation, forage and seed yield, and forage quality. We genotyped replicate pools of phenotypically extreme individuals (early and late-elongating) within cultivar Lea using genotyping-by-sequencing in pools (pool-GBS). After calling and filtering SNPs and GBS locus haplotype polymorphisms, we estimated allele frequencies and searched for markers with significantly different allele frequencies in the two phenotypic groups using BayeScan, an FST-based test utilizing replicate pools, and a test based on error variance of replicate pools. Of the three methods, BayeScan was the least stringent, and the error variance-based test the most stringent. Fifteen significant markers were identified in common by all three tests. The candidate genes flanking the markers include genes with potential roles in the vernalization, autonomous, and photoperiod regulation of floral transition, hormonal regulation of stem elongation, and cell growth. These results provide a first insight into the potential genes and mechanisms controlling transition to stem elongation in a perennial legume, which lays a foundation for further functional studies of the genetic determinants regulating this important trait.
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Trifolium , Mapeo Cromosómico/métodos , Frecuencia de los Genes , Genotipo , Polimorfismo de Nucleótido Simple/genética , Trifolium/genéticaRESUMEN
BACKGROUND: The availability of chromosome-scale genome assemblies is fundamentally important to advance genetics and breeding in crops, as well as for evolutionary and comparative genomics. The improvement of long-read sequencing technologies and the advent of optical mapping and chromosome conformation capture technologies in the last few years, significantly promoted the development of chromosome-scale genome assemblies of model plants and crop species. In grasses, chromosome-scale genome assemblies recently became available for cultivated and wild species of the Triticeae subfamily. Development of state-of-the-art genomic resources in species of the Poeae subfamily, which includes important crops like fescues and ryegrasses, is lagging behind the progress in the cereal species. RESULTS: Here, we report a new chromosome-scale genome sequence assembly for perennial ryegrass, obtained by combining PacBio long-read sequencing, Illumina short-read polishing, BioNano optical mapping and Hi-C scaffolding. More than 90% of the total genome size of perennial ryegrass (approximately 2.55 Gb) is covered by seven pseudo-chromosomes that show high levels of collinearity to the orthologous chromosomes of Triticeae species. The transposon fraction of perennial ryegrass was found to be relatively low, approximately 35% of the total genome content, which is less than half of the genome repeat content of cultivated cereal species. We predicted 54,629 high-confidence gene models, 10,287 long non-coding RNAs and a total of 8,393 short non-coding RNAs in the perennial ryegrass genome. CONCLUSIONS: The new reference genome sequence and annotation presented here are valuable resources for comparative genomic studies in grasses, as well as for breeding applications and will expedite the development of productive varieties in perennial ryegrass and related species.