Implementation of two high through-put techniques in a novel application: detecting point mutations in large EMS mutated plant populations

Background - The establishment of mutant populations together with the strategies for targeted mutation detection has been applied successfully to a large number of organisms including many species in the plant kingdom. Considerable efforts have been invested into research on tomato as a model for berry-fruit plants. With the progress of the tomato sequencing project, reverse genetics becomes an obvious and achievable goal. Results - Here we describe the treatment of Solanum lycopersicum seeds with 1% EMS and the development of a new mutated tomato population. To increase targeted mutant detection throughput an automated seed DNA extraction has been combined with novel mutation detection platforms for TILLING in plants. We have adapted two techniques used in human genetic diagnostics: Conformation Sensitive Capillary Electrophoresis (CSCE) and High Resolution DNA Melting Analysis (HRM) to mutation screening in DNA pools. Classical TILLING involves critical and time consuming steps such as endonuclease digestion reactions and gel electrophoresis runs. Using CSCE or HRM, the only step required is a simple PCR before either capillary electrophoresis or DNA melting curve analysis. Here we describe the development of a mutant tomato population, the setting up of two polymorphism detection platforms for plants and the results of the first screens as mutation density in the populations and estimation of the false-positives rate when using HRM to screen DNA pools. Conclusion - These results demonstrate that CSCE and HRM are fast, affordable and sensitive techniques for mutation detection in DNA pools and therefore allow the rapid identification of new allelic variants in a mutant population. Results from the first screens indicate that the mutagen treatment has been effective with an average mutation detection rate per diploid genome of 1.36 mutation/kb/1000 line

Genomic Architecture and Evolution of the Cellulose synthase Gene Superfamily as Revealed by Phylogenomic Analysis

The Cellulose synthase superfamily synthesizes cellulose and different hemicellulosic polysaccharides in plant cell walls. While much has been discovered about the evolution and function of these genes, their genomic architecture and relationship with gene (sub-)functionalization and evolution remains unclear. By using 242 genomes covering plant evolution from green algae to eudicots, we performed a large-scale analysis of synteny, phylogenetic, and functional data of the CesA superfamily. Results revealed considerable gene copy number variation across species and gene families, and also two patterns – singletons vs. tandem arrays – in chromosomic gene arrangement. Synteny analysis revealed exceptional conservation of gene architecture across species, but also lineage-specific patterns across gene (sub-)families. Synteny patterns correlated with gene sub-functionalization into primary and secondary CesAs and distinct CslD functional isoforms. Furthermore, a genomic context shift of a group of cotton secondary CesAs was associated with peculiar properties of cotton fiber synthesis. Finally, phylogenetics suggested that primary CesA sequences appeared before the secondary CesAs, while phylogenomic analyses unveiled the genomic trace of the CslD duplication that initiated the CslF family. Our results describe in detail the genomic architecture of the CesA superfamily in plants, highlighting its crucial relevance for gene diversification and sub-functionalization, and for understanding their evolution

Highly differentiated genomic properties underpin the different cell walls of Poaceae and eudicots

Plant cell walls of Poaceae and eudicots differ substantially, both in the content and composition of their components. However, the genomic and genetic basis underlying these differences is not fully resolved. In this research, we analyzed multiple genomic properties of 150 cell wall gene families across 169 angiosperm genomes. The properties analyzed include gene presence/absence, copy number, synteny, occurrence of tandem gene clusters, and phylogenetic gene diversity. Results revealed a profound genomic differentiation of cell wall genes between Poaceae and eudicots, often associated with the cell wall diversity between these plant groups. For example, overall patterns of gene copy number variation and synteny were clearly divergent between Poaceae and eudicot species. Moreover, differential Poaceae–eudicot copy number and genomic contexts were observed for all the genes within the BEL1-like HOMEODOMAIN 6 regulatory pathway, which respectively induces and represses secondary cell wall synthesis in Poaceae and eudicots. Similarly, divergent synteny, copy number, and phylogenetic gene diversification were observed for the major biosynthetic genes of xyloglucans, mannans, and xylans, potentially contributing to the differences in content and types of hemicellulosic polysaccharides differences in Poaceae and eudicot cell walls. Additionally, the Poaceae-specific tandem clusters and/or higher copy number of PHENYLALANINE AMMONIA-LYASE, CAFFEIC ACID O-METHYLTRANSFERASE, or PEROXIDASE genes may underly the higher content and larger variety of phenylpropanoid compounds observed in Poaceae cell walls. All these patterns are discussed in detail in this study, along with their evolutionary and biological relevance for cell wall (genomic) diversification between Poaceae and eudicots

Detection and Analysis of Syntenic Quantitative Trait Loci Controlling Cell Wall Quality in Angiosperms

Translational genomics can enable a quicker improvement of orphan crops toward novel agricultural applications, including the advancement of orphan biomass species for cultivation on marginal lands. In this sense, cell wall quality is a preeminent breeding target. However, tools to efficiently project genetic data on target traits across large sets of species are currently missing. This study aimed at closing this gap by developing a strategy to project a set of cell wall QTLs across a large group of plants by using genome synteny. This strategy is suited for large-scale analyses and detected 362 syntenic cell wall QTLs (SQTLs) across 74 angiosperms, including several (orphan) biomass species. SQTLs analyses revealed that they span large portions of the initial cell wall QTLs and are extensively conserved across diverse species. Moreover, numerous QTLs cell wall genes were conserved through SQTLs, including genes displaying allelic variation associated with cell wall composition. Functional analyses showed that highly conserved genes of SQTLs include important cell wall transcription factors and genes involved in the remodeling of cell wall polymers. For some of these gene families, SQTLs indicated the presence of differentially conserved genomic contexts for different gene members, highlighting their utility as a tool to pinpoint gene targets that maximize the likelihood of functional gene conservation. Overall, the results of this study can facilitate “universal” approaches for breeding (orphan) biomass crops, while the strategy for QTLs translation can be applied to other sets of traits and species, helping to unlock the potential of orphan species

A genome wide association study to dissect the genetic architecture of agronomic traits in Andean lupin (Lupinus mutabilis)

Establishing Lupinus mutabilis as a protein and oil crop requires improved varieties adapted to EU climates. The genetic regulation of strategic breeding traits, including plant architecture, growing cycle length and yield, is unknown. This study aimed to identify associations between 16 669 single nucleotide polymorphisms (SNPs) and 9 agronomic traits on a panel of 223 L. mutabilis accessions, grown in four environments, by applying a genome wide association study (GWAS). Seven environment-specific QTLs linked to vegetative yield, plant height, pods number and flowering time, were identified as major effect QTLs, being able to capture 6 to 20% of the phenotypic variation observed in these traits. Furthermore, two QTLs across environments were identified for flowering time on chromosome 8. The genes FAF, GAMYB and LNK, regulating major pathways involved in flowering and growth habit, as well as GA30X1, BIM1, Dr1, HDA15, HAT3, interacting with these pathways in response to hormonal and environmental cues, were prosed as candidate genes. These results are pivotal to accelerate the development of L. mutabilis varieties adapted to European cropping conditions by using marker-assisted selection (MAS), as well as to provide a framework for further functional studies on plant development and phenology in this species

Elucidating the Genetic Architecture of Fiber Quality in Hemp (Cannabis sativa L.) Using a Genome-Wide Association Study

Hemp (Cannabis sativa L.) is a bast-fiber crop with a great potential in the emerging bio-based economy. Yet, hemp breeding for fiber quality is restricted and that is mainly due to the limited knowledge of the genetic architecture of its fiber quality. A panel of 123 hemp accessions, with large phenotypic variability, was used to study the genetic basis of seven cell wall and bast fiber traits relevant to fiber quality. These traits showed large genetic variance components and high values of broad sense heritability in this hemp panel, as concluded from the phenotypic evaluation across three test locations with contrasting environments. The hemp panel was genotyped using restriction site associated DNA sequencing (RAD-seq). Subsequently, a large set (> 600,000) of selected genome-wide single nucleotide polymorphism (SNP) markers was used for a genome-wide association study (GWAS) approach to get insights into quantitative trait loci (QTLs) controlling fiber quality traits. In absence of a complete hemp genome sequence, identification of QTLs was based on the following characteristics: (i) association level to traits, (ii) fraction of explained trait variance, (iii) collinearity between QTLs, and (iv) detection across different environments. Using this approach, 16 QTLs were identified across locations for different fiber quality traits, including contents of glucose, glucuronic acid, mannose, xylose, lignin, and bast fiber content. Among them, six were found across the three environments. The genetic markers composing the QTLs that are common across locations are valuable tools to develop novel genotypes of hemp with improved fiber quality. Underneath the QTLs, 12 candidate genes were identified which are likely to be involved in the biosynthesis and modification of monosaccharides, polysaccharides, and lignin. These candidate genes were suggested to play an important role in determining fiber quality in hemp. This study provides new insights into the genetic architecture of fiber traits, identifies QTLs and candidate genes that form the basis for molecular breeding for high fiber quality hemp cultivars

Rinitis y factores psicológicos: estado de su relación (The association between rhinitis and psychological factors: an update)

The present article examines the concept of rhinitis, its classification and the reasons for classifying the different typesof rhinitis as psychophysiological diseases. We review the few studies that have investigated the association betweenrhinitis and psychological factors. Most of these studies have focussed on allergies, among which is rhinitis. We alsoassess current knowledge on the association between rhinitis and psychological factors, mainly addressing anxiety,depression and personality characteristics. Further lines of research are suggested regarding this association

High-Resolution Analysis of Growth and Transpiration of Quinoa Under Saline Conditions

The Plantarray 3.0 phenotyping platform® was used to monitor the growth and water use of the quinoa varieties Pasto and selRiobamba under salinity (0–300 mM NaCl). Salinity reduced the cumulative transpiration of both varieties by 60% at 200 mM NaCl and by 75 and 82% at 300 mM NaCl for selRiobamba and Pasto, respectively. Stomatal conductance was reduced by salinity, but at 200 mM NaCl Pasto showed a lower reduction (15%) than selRiobamba (35%), along with decreased specific leaf area. Diurnal changes in water use parameters indicate that under salt stress, daily transpiration in quinoa is less responsive to changes in light irradiance, and stomatal conductance is modulated to maximize CO2 uptake and minimize water loss following the changes in VPD (vapor pressure deficit). These changes might contribute to the enhanced water use efficiency of both varieties under salt stress. The mechanistic crop model LINTUL was used to integrate physiological responses into the radiation use efficiency of the plants (RUE), which was more reduced in Pasto than selRiobamba under salinity. By the end of the experiment (eleven weeks after sowing, six weeks after stress), the growth of Pasto was significantly lower than selRiobamba, fresh biomass was 50 and 35% reduced at 200 mM and 70 and 50% reduced at 300 mM NaCl for Pasto and selRiobamba, respectively. We argue that contrasting water management strategies can at least partly explain the differences in salt tolerance between Pasto and selRiobamba. Pasto adopted a “conservative-growth” strategy, saving water at the expense of growth, while selRiobamba used an “acquisitive-growth” strategy, maximizing growth in spite of the stress. The implementation of high-resolution phenotyping could help to dissect these complex growth traits that might be novel breeding targets for abiotic stress tolerance

Differential responses to salt stress in ion dynamics, growth and seed yield of European quinoa varieties

Quinoa is a nutritious seed crop with a great potential to grow in saline soils. Here, we studied ion concentrations in quinoa tissues throughout the life cycle of the plant, and linked ion dynamics to responses in growth parameters, seed yield and efficiency of photosynthesis under salinity (0–400 mM NaCl). Ion dynamics changed from high ion exclusion (>99 %, root contents lower than root medium and low accumulation of ions in the leaves) before flowering, to a build-up of ions during seed filling. This indicates a change in strategy in maintaining the necessary gradient of water potential from the root medium to the leaves. K+ concentrations in leaves also increased by more than 100 % in response to prolonged severe salt stress, which may point to a role of this ion in leaf osmotic adjustment. Accumulation of ions in epidermal bladder cells did not contribute substantially to Na+-exclusion as it was less than 6 % of the total Na+ taken up in leaves. Growth under salt stress was mostly impaired by anatomical adaptations (reduced SLA), while initial light use efficiency (Fv/Fm) and NAR were not affected. The variety Pasto showed a “survival strategy” to high salinity with higher ion exclusion and a higher reduction in transpiration than the other varieties, at the expense of lower biomass and seed yield.</p