Towards a comprehensive characterization of durum wheat landraces in Moroccan traditional agrosystems: analysing genetic diversity in the light of geography, farmers’ taxonomy and tetraploid wheat domestication history

Background: Crop diversity managed by smallholder farmers in traditional agrosystems is the outcome of historical and current processes interacting at various spatial scales, and influenced by factors such as farming practices and environmental pressures. Only recently have studies started to consider the complexity of these processes instead of simply describing diversity for breeding purposes. A first step in that aim is to add multiple references to the collection of genetic data, including the farmers' varietal taxonomy and practices and the historical background of the crop. Results: On the basis of interview data collected in a previous study, we sampled 166 populations of durum wheat varieties in two traditional Moroccan agrosystems, in the Pre-Rif and Atlas Mountains regions. Using a common garden experiment, we detected a high phenotypic variability on traits indicative of taxonomical position and breeding status, namely spike shape and plant height. Populations often combined modern (short) with traditional-like (tall) statures, and classical durum squared spike shape (5 flowers/spikelet) with flat spike shape (3 flowers/spikelet) representative of primitive domesticated tetraploid wheat (ssp. dicoccum). By contrast, the genetic diversity assessed using 14 microsatellite markers was relatively limited. When compared to the genetic diversity found in a large collection of tetraploid wheat, it corresponded to free-threshing tetraploid wheat. Within Morocco, the two studied regions differed for both genetic diversity and variety names. Within regions, neither geography nor variety names nor even breeding status constituted strong barriers to gene exchange despite a few significant patterns. Conclusions: This first assessment of morphological and genetic diversity allowed pointing out some important factors that may have influenced the structure and evolutionary dynamics of durum wheat in Morocco: the significance of variety names, the occurrence of mixtures within populations, the relative strength of seed exchange between farmers and local adaptation, as well as the fate of modern varieties once they have been introduced. Further, multidisciplinary studies at different spatial scales are needed to better understand these complex agrosystems of invaluable importance for food security

A small XY chromosomal region explains sex determination in wild dioecious V. vinifera and the reversal to hermaphroditism in domesticated grapevines

Publis014-agap-029Background In Vitis vinifera L., domestication induced a dramatic change in flower morphology: the wild sylvestris subspecies is dioecious while hermaphroditism is largely predominant in the domesticated subsp. V. v. vinifera. The characterisation of polymorphisms in genes underlying the sex-determining chromosomal region may help clarify the history of domestication in grapevine and the evolution of sex chromosomes in plants. In the genus Vitis, sex determination is putatively controlled by one major locus with three alleles, male M, hermaphrodite H and female F, with an allelic dominance M > H > F. Previous genetic studies located the sex locus on chromosome 2. We used DNA polymorphisms of geographically diverse V. vinifera genotypes to confirm the position of this locus, to characterise the genetic diversity and traces of selection in candidate genes, and to explore the origin of hermaphroditism. Results In V. v. sylvestris, a sex-determining region of 154.8 kb, also present in other Vitis species, spans less than 1% of chromosome 2. It displays haplotype diversity, linkage disequilibrium and differentiation that typically correspond to a small XY sex-determining region with XY males and XX females. In male alleles, traces of purifying selection were found for a trehalose phosphatase, an exostosin and a WRKY transcription factor, with strikingly low polymorphism levels between distant geographic regions. Both diversity and network analysis revealed that H alleles are more closely related to M than to F alleles. Conclusions Hermaphrodite alleles appear to derive from male alleles of wild grapevines, with successive recombination events allowing import of diversity from the X into the Y chromosomal region and slowing down the expansion of the region into a full heteromorphic chromosome. Our data are consistent with multiple domestication events and show traces of introgression from other Asian Vitis species into the cultivated grapevine gene pool. La vigne domestiquée, Vitis vinifera ssp. vinifera, cultivée pour la production de fruit et de vin à travers le monde, dérive de la vigne sauvage, Vitis vinifera ssp. sylvestris, sous-espèce endémique de l'Eurasie. Au cours de la domestication, le système de reproduction a évolué de la diécie à l'hermaphrodisme. Nous montrons que la région du sexe est sous le contrôle d'une région chromosomique qui couvre au maximum 154kpbp, moins de 1% du chromosome 2. La caractérisation de ce locus en terme de diversité haplotypique, de signature de sélection et de déséquilibre de liaison a permis de révéler un système de détermination sexuelle de type XY. La petite taille de cette région chromosomique semble indiquer un stade très précoce dans l'évolution de chromosomes sexuels, malgré que la diécie soit le trait ancestral chez toutes les espèces de Vitis, ayant divergé du sous-genre Muscadinia il y a plusieurs millions d'années. L'analyse des distances génétiques entre haplotypes dans le locus du sexe a révélé que l'hermaphrodisme observé chez la vigne domestiquée résulte de la mutation de l'allèle mâle présent chez la vigne sauvage. Le réseau d'haplotypes a montré qu'en plus de la contribution de V. sylvestris, une autre espèce de Vitis asiatique a pu contribuer à la constitution du génome actuel de la vigne cultivée moderne. Ces travaux résultent d'une collaboration entre l'équipe DAAV d'AGAP et l'UMR CBAE (Montpellier)

Histoire de la domestication de Triticum turgidum : la capture d'exons au service de l'étude de la diversité génétique

Depuis le début de sa domestication dans le croissant fertile, il y a 12 000 ans, le blé dur, Triticum turgidum a subi de nombreux évènements démographiques et sélectifs. La première transition caractérisant le passage du blé dur sauvage (amidonnier sauvage) Triticum turgidum spp dicoccoïdes, à la première forme cultivée (amidonnier cultivé) Triticum turgidum spp dicoccum est marquée, entre autre, par l’apparition d’un rachis solide. Ce caractère, contrôlé notamment par les gènes Br, a permis aux premiers agriculteurs de récolter, sur les plantes, le grain à maturité. La deuxième transition s’est produite entre l’amidonnier cultivé et Triticum turgidum spp durum. Elle est caractérisée par l’apparition des grains nus permettant un battage plus facile (caractère contrôlé notamment par le gène Q). La dernière transition majeure intervient lors de la révolution verte, dans les années 1960, et marque le passage des « populations de pays » aux variétés « élites », sélectionnées pour de nombreux caractères morphologiques comme une taille réduite (gène Rht), un rendement plus fort ou une capacité plus grande à absorber beaucoup d’azote sans que les plantes ne soient sujettes à la verse. Dans le but d’affiner nos connaissances sur l’histoire évolutive de l’espèce Triticum turgidum, nous avons produit un jeu de données moléculaires en utilisant la méthode d’enrichissement par capture (10 000 régions de 120pb situées dans la partie codante du génome) pour contourner les difficultés liées à la grande taille de son génome (10.5 Gb). Cette étude a été réalisée sur 120 génotypes, 30 pour chacune des quatre formes évolutives précédemment citées. L’analyse de la diversité nucléotidique obtenue (135 863 SNPs) nous a permis d’estimer la réduction de celle-ci associée à chaque transition. Le goulot d’étranglement le plus important a eu lieu lors du premier évènement de domestication avec seulement 59 % de la diversité présente chez Triticum turgidum spp dicoccoïdes encore présente chez la forme cultivée, Triticum turgidum spp dicoccum. Pour la deuxième transition, nous avons estimé que 76% de la diversité de Triticum turgidum spp dicoccum se retrouvait dans le groupe Triticum turgidum ssp durum. Et pour finir, 91% de la diversité est conservée entre les deux formes de T. turgidum ssp durum. L’utilisation de la séquence génomique de référence (accession «Zavitan» - Avni et al. 2017; Zhu et al. 2019), nous a également permis d’observer la variation du niveau de diversité le long du génome pour chacune des quatre formes évolutives. Nous avons observé que certaines zones ont vu leur niveau de diversité considérablement diminuer au cours de la domestication, alors que d’autres sont encore très polymorphes. Nous avons également pu détecter des signatures de sélection, sur ces quatre formes, d’abord sans a priori, puis en ciblant les locus impliqués dans l’expression de traits phénotypiques caractéristiques de la domestication (locus Br, Q et Rht) ainsi que deux QTLs impliqués dans le poids des grains et la teneur en azote dans la feuille, trait qui reflète la stratégie d’acquisition des ressources de la plante. Ce travail a permis de proposer de nouvelles perspectives pour le développement d’outils moléculaires et des nouvelles pistes pour affiner notre compréhension de l’histoire évolutive de l’espèce Triticum turgidum

Versatile and efficient RNA extraction protocol for grapevine berry tissue, suited for next generation RNA sequencing

Background and Aims: New high-throughput tools for transcriptomic analysis such as RNA sequencing have been developed rapidly in recent years. These technologies provide new opportunities for biologists to improve the understanding of gene expression underlying important physiological processes. The sequencing of RNA by nextgeneration technologies is, in particular, dependent on the use of pure and un-degraded RNA. Many fruits from perennial crops share an elevated concentration of tannins and polysaccharides and a strong dilution of cytosolic content upon vacuole hypertrophy. The grapevine berry presents these characteristics together with high vacuole acidity, which has never been taken into account in previous protocols, and poses thereby a particular challenge for RNA extraction. Methods and Results: In order to counterbalance these limitations, we developed a new extraction protocol based on a tri-sodium-citrate extraction buffer that has a high buffer capacity and proved to be particularly suitable for low pH plant tissue. Conclusions: The proposed RNA extraction procedure produces consistently high-quality RNA (RNA integrity number > 8 up to 10; 260 nm/280 nm > 2; 260 nm/230 nm > 1.9) with an elevated yield [20 μg/g fresh mass (FM) for ripe berries up to 150 μg/FM for green berries]. The method was developed initially for berry tissues but proved also to be efficient for other grapevine tissues, such as nodes, roots, leaves, seeds, lignified shoots and flowers. Significance of the Study: The method is most suitable for modern gene expression analysis methods, such as RNA sequencing and microarray studies. Successful construction of cDNA libraries and high numbers of detected reads obtained by next-generation RNA sequencing underline the applicability of the protocol

Elite durum wheat genetic map and recombination rate variation in a multiparental connected design

UMR 1334 AGAP : Equipe AFEF ‘Architecture et Fonctionnement des Espèces fruitières’ ; Team AFFS ‘Architecture and Functioning of Fruit Species’ Contact : [email protected] maps published for elite durum wheat (Triticum turgidum durum Desf.), were mainly related to Italian and North American pools. To provide additional information dedicated to durum elite pool, a recombinant inbred population (217 RILs) derived form 2 French commercial varieties (Ixos and Primadur), has been mapped with 529 loci (98 AFLPs, 67 SSRs, 1 ISBP and 363 DArTs). Genetic map spanned a total of 2,082 cM (5.7 cM/marker on average). Recombination rate variation over the genome was documented through the analysis of six segregating populations representing a total of 48 RILs x 6, issued from an half diallel design based on four elite lines-Neodur, Ixos, Lloyd and Primadur-including the 2 previous genitors. Each set of three genetically connected populations (48 RILs x 3) was used to build the consensus parental maps for each genitor. To construct the four consensus parental maps, marker ordering was determined on 217 RILs map which was an extended population of the most polymorphic cross (Ixos x Primadur). Sizeable, recombination rate variation has been observed between the four parental maps both at the marker pair scale and at the linkage group scale. Two of the parents, Lloyd and Primadur, had strong, opposite effects increasing and decreasing, respectively the recombination rate. Additional studies are proposed to increase our understanding of this variation, with the identification of environmental or genetic factors affecting recombination rate

Low growth temperatures uncouple sugar accumulation from malic acid degradation in ripenning microvine berries

International audienc

Genotyping by targeted sequence capture in durum wheat: Application to detect QTL for wheat spindle streak mosaic virus resistance

International audienceRecent advances in genotyping and phenotyping technologies can now be applied in durum wheat and permit to adress a genome wide approach to disclose the geneti c architecture of kernel quality traits. DNA chips can allow the genotyping of thousands of SNP markers on several hundreds of lines while NIRS (Near Infra Red spectroscopy) allow the predicti on of relevant traits in durum wheat quality, e.g., protein content, yellow pigment content and semolina yield. Genome Wide Assocati on Studies (GWAS) have proven their potenti al in detecti ng chromosomal areas implied in the variati on of traits. Good panels should have a diversity level, a low structurati on and reduced level of linkage disequilibrium. Assembling panels from lines of diverse origin is usually the rule in many crops but may lead to a substanti al structurati on eff ect and to spurrious associati ons. We propose here to use lines drawn from an original composite cross populati on, originally dedicated to create a pre breeding resource. The Evoluti onary Prebreeding pOpulati on (EPO) is conti nuously grown since 1997 with a light selecti on and a 10% outcrossing rate. We report the fi rst results of a GWAS study on 300 hundred lines drawn from the 2009 harvest of EPO. Using a set of 62 000 polymorphic SNP drawn along the genome, we report the associati ons found between this set of markers and a set of traits related to the kernel quality predicted by NIRS. First, the characteristi cs of the EPO populati on will be illustrated to examine its power to detect associati on. We will report the fi ne geneti c structurati on and the fi ne scale Linkage disequilibrium on the 3B chromosome on which the SNP can be physically located. Then kernels quality traits will be dissected by GWAS to detect the most important chromosomal regions involved in the variati on. Finally, each trait will be predicted as the whole level to discuss the potenti al of Genomic Selecti on (GS) in the improvement of quality in the durum kernel

Temperature desynchronizes sugar and organic acid metabolism in ripening grapevine fruits and remodels their transcriptome

Fruit composition at harvest is strongly dependent on the temperature during the grapevine developmental cycle. This raises serious concerns regarding the sustainability of viticulture and the socio-economic repercussions of global warming for many regions where the most heat-tolerant varieties are already cultivated. Despite recent progress, the direct and indirect effects of temperature on fruit development are far from being understood. Experimental limitations such as fluctuating environmental conditions, intra-cluster heterogeneity and the annual reproductive cycle introduce unquantifiable biases for gene expression and physiological studies with grapevine. In the present study, DRCF grapevine mutants (microvine) were grown under several temperature regimes in duly-controlled environmental conditions. A singly berry selection increased the accuracy of fruit phenotyping and subsequent gene expression analyses. The physiological and transcriptomic responses of five key stages sampled simultaneously at day and nighttime were studied by RNA-seq analysis. A total of 674 millions reads were sequenced from all experiments. Analysis of differential expression yielded in a total of 10 788 transcripts modulated by temperature. An acceleration of green berry development under higher temperature was correlated with the induction of several candidate genes linked to cell expansion. High temperatures impaired tannin synthesis and degree of galloylation at the transcriptomic levels. The timing of malate breakdown was delayed to mid-ripening in transgressively cool conditions, revealing unsuspected plasticity of berry primary metabolism. Specific ATPases and malate transporters displayed development and temperature-dependent expression patterns, besides less marked but significant regulation of other genes in the malate pathway. The present study represents, to our knowledge the first abiotic stress study performed on a fleshy fruits model using RNA-seq for transcriptomic analysis. It confirms that a careful stage selection and a rigorous control of environmental conditions are needed to address the long-term plasticity of berry development with respect to temperature. Original results revealed temperature-dependent regulation of key metabolic processes in the elaboration of berry composition. Malate breakdown no longer appears as an integral part of the veraison program, but as possibly triggered by an imbalance in cytoplasmic sugar, when efficient vacuolar storage is set on with ripening, in usual temperature conditions. Furthermore, variations in heat shock responsive genes that will be very valuable for further research on temperature adaptation of plants have been evidenced