Caractérisation de la variation phénotypique de la taille de la baie chez la vigne Vitis vinifera L. et approches de génétique d'association et de recherche de traces de sélection pour ce caractère

La taille du fruit est un caractère agronomique important pour le rendement des espèces cultivées.Or, excepté chez la tomate, les mécanismes impliqués dans la variation de la taille des fruits charnus sont encore mal connus. L'objectif de ma thèse était d'identifier les principaux facteurs anatomiques impliqués dans la variation de la taille des baies, de réaliser des tests d'association génétique sur des gènes candidats et de rechercher des possibles traces de domestication liées à la variation de la taille des baies. Le nombre de cellules déjà déterminé dans l ovaire avant l anthèse et la capacité des cellules du mésocarpe à grossir après l anthèse se sont révélés les principaux déterminants de la variation de la baie entre les variétés. Trois gènes potentiellement impliqués dans ce caractère ont été identifiés par des approches de génétique d'association. La MADS boxVvAGL11 est associée au nombre de pépins de la baie. VvHMGR1, une HMGR impliquée dans la synthèse des isoprénoïdes précurseurs de régulateurs de croissance, est associée à la longueur des baies. Enfin, VvJMJ un facteur de transcription de type jumonji est à la fois associé à la variation du poids et à la longueur des baies. La caractérisation du locus Fleshless berry (Flb) a également été poursuivie avec (i) l analyse du polymorphisme de séquence de 69 fragments de gènes entre les vignes sauvages et cultivées, ce qui a permis l identification d un gène présentant une trace de sélection et une diversité nucléotidique plus élevée dans le compartiment sauvage que dans lecultivé. (ii) La mise en évidence d une homozygotie complète d une région de 1 Mb contenant le locus Flb chez certains cultivars comme l Ugni Blanc, rendant impossible le clonage positionnel du gène dans la population F1. L ensemble de ce travail a permis de mettre en évidence la complexité des mécanismes à l origine de la variation de la taille de la baie chez la vigne. Nous avons montré la faisabilité des études de génétique d association chez la vigne en utilisant des gènes candidats pour le poids de baies, et identifié les candidats les plus intéressants pour la sélection assistée par marqueurs et les études fonctionnelles ultérieures.Fruit size is an important trait in fruit crops. However, excepted in tomato, little is known about the genetic and molecular control of fleshy fruit size variation. The aim of my thesis was to identify themain anatomical factors underlying berry size variation, to carry out an association genetics based approach with candidate genes and to search for possible signatures of domestication linked toberry size variation. Cell divisions before anthesis and cell expansion after anthesis were found tobe the major determinants of flesh weight variation between varieties in grapevine. Three geneswere identified by association genetics approaches. The VvAGL11 MADS box was found associated only with the seed number per berry. VvHMGR1, a putative HMGR involved in the synthesis of isoprenoid, a precursor of plant growth regulators, was found associated with berry length. Finally,VvJMJ encoding a putative transcription factor jumonji was associated both with berry weight andberry length variation. The characterization of the Fleshless berry locus (Flb) was also pursued with(i) the analysis of the sequence polymorphism of 69 regions between wild and cultivated grapevines,that leaded to the identification of one gene showing a trace of selection and a higher nucleotide diversity in the wild compartment than in the cultivated compartment. (ii) The discovery of acomplete homozygosity of a 1 Mb region containing the Flb locus in some cultivars such as theUgni Blanc, making impossible the fine mapping of the Flb locus in a F1 population. This study has highlighted the complexity of the mechanisms underlying the berry size variation in grapevines. We demonstrated the feasibility to screen in grapevine by association genetics candidate genes for berryweight and to select the most interesting ones for marker-assisted selection and for furtherfunctional studies.EVRY-Bib. électronique (912289901) / SudocSudocFranceF

Usefulness of two SCAR markers for marker-assisted selection of seedless grapevine cultivars

A PCR-specific marker, SCP18, was developed from a RAPD marker linked to a major locus involved in seedlessness, sdI. A preliminary study of the usefulness of SCP18 and SCC8 (a marker linked to sdI previously developed by LAHOGUE et al. 1998) for the marker assisted selection of seedless varieties was realized using various strategies: a posteriori test in seedless x seedless and seeded x seeded progenies and test of their allelic diversity in a set of 81 seedless and seeded varieties. In contrast to SCP18, SSC8 was found to be a useful marker at least in the seedless x seedless progenies and to show a good linkage disequilibrium with seedlessness in our set of varieties

Génomique comparative entre Muscadinia rotundifolia et Vitis vinifera pour faciliter l'identification de gènes de résistance

Muscadinia rotundifolia est une espèce de la famille des Vitaceae. C est un sous-genre du genre Vitis, le deuxième sous-genre étant celui des Euvitis qui comprend l espèce cultivée Vitis vinifera (2n=38). M. rotundifolia (2n=40) est une source de résistance aux maladies très importante pour l amélioration de la vigne. Son génome commence seulement à être décrit avec deux cartes génétiques récemment publiées. Ma thèse a consisté à utiliser des ressources génomiques chez M. rotundifolia cv Regale (banque BAC, collection de séquence d extrémités de BAC ou BES et séquences de BACs) pour caractériser le génome de cette espèce en comparaison avec celui de V. vinifera. Les résultats obtenus ne montrent pas de différence importante entre les génomes des deux espèces en termes de composition du génome en bases (GC%), en séquences codantes ou en éléments répétés. De même, à une échelle globale, la famille de gènes NBS-LRR semble être similaire en termes de nombre et de balance entre les sous-familles. A une échelle plus fine cependant (carte physique et séquences de BAC), des remaniements relativement importants sont observés dans des régions portant cette famille de gènes, aboutissant parfois à des contenus différents en gènes, de région normalement homologues: duplication différentielles de gènes, présence/absence de gènes.Muscadinia Rotundifolia is a species of the Vitaceae family. It is a sub-genus of the Vitis genus along with the Euvitis sub-genus, which the cultivated species Vitis vinifera belongs to. M. rotundifolia (2n=40) is a very important source of resistance to diseases in grapevine breeding programs. Its genome is only starting to be described with the recent publication of two genetic maps. The present study aimed at using M. rotundifolia cv Regale genomic resources (BAC library, BAC end sequences or BES, BAC sequences) in order to characterize the genome of this species in comparison with the genome of V. vinifera. The results showed that there is no striking difference between the two species in term of base composition (GC %), repeats frequency and gene space. The NBS LRR gene family also seems to be globally quite similar between the two species in terms of numbers and balance between subfamilies. At a finer scale (physical map and BAC sequence), frequent rearrangements are observed in genomic regions carrying the NBS-LRR gene family sometimes clearly associated with a different gene content between the two species in homologous regions: differential gene duplication, presence/absence of genes.EVRY-Bib. électronique (912289901) / SudocSudocFranceF

Colour variation in red grapevines (Vitis vinifera L.): genomic organisation, expression of flavonoid 3'-hydroxylase, flavonoid 3',5'-hydroxylase genes and related metabolite profiling of red cyanidin-/blue delphinidin-based anthocyanins in berry skin

BACKGROUND: Structural genes of the phenyl-propanoid pathway which encode flavonoid 3'- and 3',5'-hydroxylases (F3'H and F3'5'H) have long been invoked to explain the biosynthesis of cyanidin- and delphinidin-based anthocyanin pigments in the so-called red cultivars of grapevine. The relative proportion of the two types of anthocyanins is largely under genetic control and determines the colour variation among red/purple/blue berry grape varieties and their corresponding wines. RESULTS: Gene fragments of VvF3'H and VvF3'5'H, that were isolated from Vitis vinifera 'Cabernet Sauvignon' using degenerate primers designed on plant homologous genes, translated into 313 and 239 amino acid protein fragments, respectively, with up to 76% and 82% identity to plant CYP75 cytochrome P450 monooxygenases. Putative function was assigned on the basis of sequence homology, expression profiling and its correlation with metabolite accumulation at ten different ripening stages. At the onset of colour transition, transcriptional induction of VvF3'H and VvF3'5'H was temporally coordinated with the beginning of anthocyanin biosynthesis, the expression being 2-fold and 50-fold higher, respectively, in red berries versus green berries. The peak of VvF3'5'H expression was observed two weeks later concomitantly with the increase of the ratio of delphinidin-/cyanidin-derivatives. The analysis of structural genomics revealed that two copies of VvF3'H are physically linked on linkage group no. 17 and several copies of VvF3'5'H are tightly clustered and embedded into a segmental duplication on linkage group no. 6, unveiling a high complexity when compared to other plant flavonoid hydroxylase genes known so far, mostly in ornamentals. CONCLUSION: We have shown that genes encoding flavonoid 3'- and 3',5'-hydroxylases are expressed in any tissues of the grape plant that accumulate flavonoids and, particularly, in skin of ripening red berries that synthesise mostly anthocyanins. The correlation between transcript profiles and the kinetics of accumulation of red/cyanidin- and blue/delphinidin-based anthocyanins indicated that VvF3'H and VvF3'5'H expression is consistent with the chromatic evolution of ripening bunches. Local physical maps constructed around the VvF3'H and VvF3'5'H loci should help facilitate the identification of the regulatory elements of each isoform and the future manipulation of grapevine and wine colour through agronomical, environmental and biotechnological tools

Construction of nested genetic core collections to optimize the exploitation of natural diversity in Vitis vinifera L. subsp. sativa

Background: The first high quality draft of the grape genome sequence has just been published. This is a critical step in accessing all the genes of this species and increases the chances of exploiting the natural genetic diversity through association genetics. However, our basic knowledge of the extent of allelic variation within the species is still not sufficient. Towards this goal, we constructed nested genetic core collections (G-cores) to capture the simple sequence repeat (SSR) diversity of the grape cultivated compartment (Vitis vinifera L. subsp. sativa) from the world's largest germplasm collection (Domaine de Vassal, INRA Hérault, France), containing 2262 unique genotypes. Results: Sub-samples of 12, 24, 48 and 92 varieties of V. vinifera L. were selected based on their genotypes for 20 SSR markers using the M-strategy. They represent respectively 58%, 73%, 83% and 100% of total SSR diversity. The capture of allelic diversity was analyzed by sequencing three genes scattered throughout the genome on 233 individuals: 41 single nucleotide polymorphisms (SNPs) were identified using the G-92 core (one SNP for every 49 nucleotides) while only 25 were observed using a larger sample of 141 individuals selected on the basis of 50 morphological traits, thus demonstrating the reliability of the approach. Conclusion: The G-12 and G-24 core-collections displayed respectively 78% and 88% of the SNPs respectively, and are therefore of great interest for SNP discovery studies. Furthermore, the nested genetic core collections satisfactorily reflected the geographic and the genetic diversity of grape, which are also of great interest for the study of gene evolution in this species

The grapevine gene nomenclature system

[Background] Grapevine (Vitis vinifera L.) is one of the most important fruit crops in the world and serves as a valuable model for fruit development in woody species. A major breakthrough in grapevine genomics was achieved in 2007 with the sequencing of the Vitis vinifera cv. PN40024 genome. Subsequently, data on structural and functional characterization of grape genes accumulated exponentially. To better exploit the results obtained by the international community, we think that a coordinated nomenclature for gene naming in species with sequenced genomes is essential. It will pave the way for the accumulation of functional data that will enable effective scientific discussion and discovery. The exploitation of data that were generated independently of the genome release is hampered by their heterogeneous nature and by often incompatible and decentralized storage. Classically, large amounts of data describing gene functions are only available in printed articles and therefore remain hardly accessible for automatic text mining. On the other hand, high throughput >Omics> data are typically stored in public repositories, but should be arranged in compendia to better contribute to the annotation and functional characterization of the genes.[Results] With the objective of providing a high quality and highly accessible annotation of grapevine genes, the International Grapevine Genome Project (IGGP) commissioned an international Super-Nomenclature Committee for Grape Gene Annotation (sNCGGa) to coordinate the effort of experts to annotate the grapevine genes. The goal of the committee is to provide a standard nomenclature for locus identifiers and to define conventions for a gene naming system in this paper.[Conclusions] Learning from similar initiatives in other plant species such as Arabidopsis, rice and tomato, a versatile nomenclature system has been developed in anticipation of future genomic developments and annotation issues. The sNCGGa's first outreach to the grape community has been focused on implementing recommended guidelines for the expert annotators by: (i) providing a common annotation platform that enables community-based gene curation, (ii) developing a gene nomenclature scheme reflecting the biological features of gene products that is consistent with that used in other organisms in order to facilitate comparative analyses. © 2014 Grimplet et al.Authors would like to thank the Grape Research Coordination Network (NSF grant DBI 0741876) for financial support, the International Grape Genome Program and the COST action FA1106 “Quality fruit”. J.G. was supported by the Ramon y Cajal program (RYC-2011-07791). We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI).Peer Reviewe

Molecular, genetic and transcriptional evidence for a role of VvAGL11 in stenospermocarpic seedlessness in grapevine

<p>Abstract</p> <p>Background</p> <p>Stenospermocarpy is a mechanism through which certain genotypes of <it>Vitis vinifera </it>L. such as Sultanina produce berries with seeds reduced in size. Stenospermocarpy has not yet been characterized at the molecular level.</p> <p>Results</p> <p>Genetic and physical maps were integrated with the public genomic sequence of <it>Vitis vinifera </it>L. to improve QTL analysis for seedlessness and berry size in experimental progeny derived from a cross of two seedless genotypes. Major QTLs co-positioning for both traits on chromosome 18 defined a 92-kb confidence interval. Functional information from model species including <it>Vitis </it>suggested that <it>VvAGL11</it>, included in this confidence interval, might be the main positional candidate gene responsible for seed and berry development.</p> <p>Characterization of <it>VvAGL11 </it>at the sequence level in the experimental progeny identified several SNPs and INDELs in both regulatory and coding regions. In association analyses performed over three seasons, these SNPs and INDELs explained up to 78% and 44% of the phenotypic variation in seed and berry weight, respectively. Moreover, genetic experiments indicated that the regulatory region has a larger effect on the phenotype than the coding region. Transcriptional analysis lent additional support to the putative role of <it>VvAGL11's </it>regulatory region, as its expression is abolished in seedless genotypes at key stages of seed development. These results transform <it>VvAGL11 </it>into a functional candidate gene for further analyses based on genetic transformation.</p> <p>For breeding purposes, intragenic markers were tested individually for marker assisted selection, and the best markers were those closest to the transcription start site.</p> <p>Conclusion</p> <p>We propose that <it>VvAGL11 </it>is the major functional candidate gene for seedlessness, and we provide experimental evidence suggesting that the seedless phenotype might be caused by variations in its promoter region. Current knowledge of the function of its orthologous genes, its expression profile in <it>Vitis </it>varieties and the strong association between its sequence variation and the degree of seedlessness together indicate that the D-lineage MADS-box gene <it>VvAGL11 </it>corresponds to the <it>Seed Development Inhibitor locus </it>described earlier as a major locus for seedlessness. These results provide new hypotheses for further investigations of the molecular mechanisms involved in seed and berry development.</p

Towards an Open Grapevine Information System

Viticulture, like other fields of agriculture, is currently facing important challenges that will be addressed only through sustained, dedicated and coordinated research. Although the methods used in biology have evolved tremendously in recent years and now involve the routine production of large data sets of varied nature, in many domains of study, including grapevine research, there is a need to improve the findability, accessibility, interoperability and reusability (FAIR-ness) of these data. Considering the heterogeneous nature of the data produced, the transnational nature of the scientific community and the experience gained elsewhere, we have formed an open working group, in the framework of the International Grapevine Genome Program (www.vitaceae.org), to construct a coordinated federation of information systems holding grapevine data distributed around the world, providing an integrated set of interfaces supporting advanced data modeling, rich semantic integration and the next generation of data mining tools. To achieve this goal, it will be critical to develop, implement and adopt appropriate standards for data annotation and formatting. The development of this system, the GrapeIS, linking genotypes to phenotypes, and scientific research to agronomical and oeneological data, should provide new insights into grape biology, and allow the development of new varieties to meet the challenges of biotic and abiotic stress, environmental change, and consumer demand

A physical map of the heterozygous grapevine 'Cabernet Sauvignon' allows mapping candidate genes for disease resistance

<p>Abstract</p> <p>Background</p> <p>Whole-genome physical maps facilitate genome sequencing, sequence assembly, mapping of candidate genes, and the design of targeted genetic markers. An automated protocol was used to construct a <it>Vitis vinifera </it>'Cabernet Sauvignon' physical map. The quality of the result was addressed with regard to the effect of high heterozygosity on the accuracy of contig assembly. Its usefulness for the genome-wide mapping of genes for disease resistance, which is an important trait for grapevine, was then assessed.</p> <p>Results</p> <p>The physical map included 29,727 BAC clones assembled into 1,770 contigs, spanning 715,684 kbp, and corresponding to 1.5-fold the genome size. Map inflation was due to high heterozygosity, which caused either the separation of allelic BACs in two different contigs, or local mis-assembly in contigs containing BACs from the two haplotypes. Genetic markers anchored 395 contigs or 255,476 kbp to chromosomes. The fully automated assembly and anchorage procedures were validated by BAC-by-BAC blast of the end sequences against the grape genome sequence, unveiling 7.3% of chimerical contigs. The distribution across the physical map of candidate genes for non-host and host resistance, and for defence signalling pathways was then studied. NBS-LRR and RLK genes for host resistance were found in 424 contigs, 133 of them (32%) were assigned to chromosomes, on which they are mostly organised in clusters. Non-host and defence signalling genes were found in 99 contigs dispersed without a discernable pattern across the genome.</p> <p>Conclusion</p> <p>Despite some limitations that interfere with the correct assembly of heterozygous clones into contigs, the 'Cabernet Sauvignon' physical map is a useful and reliable intermediary step between a genetic map and the genome sequence. This tool was successfully exploited for a quick mapping of complex families of genes, and it strengthened previous clues of co-localisation of major NBS-LRR clusters and disease resistance <it>loci </it>in grapevine.</p