In order to investigate the comparability of microsatellite profiles obtained in different laboratories, ten partners in seven countries analyzed 46 grape cultivars at six loci (VVMD5, VVMD7, VVMD27, VVS2, VrZAG62, and VrZAG79). No effort was made to standardize equipment or protocols. Although some partners obtained very similar results, in other cases different absolute allele sizes and, sometimes, different relative allele sizes were obtained. A strategy for data comparison by means of reference to the alleles detected in well-known cultivars was proposed. For each marker, each allele was designated by a code based on the name of the reference cultivar carrying that allele. Thirty-three cultivars, representing from 13 to 23 alleles per marker, were chosen as references. After the raw data obtained by the different partners were coded, more than 97% of the data were in agreement. Minor discrepancies were attributed to errors, suboptimal amplification and visualization, and misscoring of heterozygous versus homozygous allele pairs. We have shown that coded microsatellite data produced in different laboratories with different protocols and conditions can be compared, and that it is suitable for the identification and SSR allele characterization of cultivars. It is proposed that the six markers employed here, already widely used, be adopted as a minimal standard marker set for future grapevine cultivar analyses, and that additional cultivars be characterized by means of the coded reference alleles presented here. The complete database is available at http://www.genres.de/eccdb/vitis/ Cuttings of the 33 reference cultivars are available on request from the Institut National de la Recherche Agronomique Vassal collection (didier.vares@ensam.inra.fr).
In this work, 78 hazelnut (Corylus avellana L.) cultivars from various germplasm repositories were studied at 16 simple sequence repeat (SSR) loci in order to identify the genotypes and investigate their genetic relations. Polymorphism at SSR loci was evaluated on the basis of number of alleles (mean: 9.4), expected heterozygosity (mean: 0.78), and power of discrimination (mean: 0.91). Several synonyms reported in the literature were confirmed, and new cases of synonymy were identified. The parentage of North American cultivars 'Butler', 'Ennis', and 'Royal', the French selection 'Fercoril-Corabel', and 'Impératrice Eugenie' was investigated on the basis of the alleles present at 16 loci and analysis at 8 additional loci. A dendrogram generated from cluster analysis using the unweighted pair group method with arithmetic mean grouped cultivars according to their pedigrees or geographical origins. There was an evident differentiation of the northern European cultivars from the southern European ones and from the Turkish cultivars. The latter clustered close to but separate from the Italian and Spanish clusters. It is very likely that exchanges of cultivars occurred between the central and western Mediterranean basin as a result of human migration and trade. A database containing the SSR profiles of the most important hazelnut cultivars will be useful for identification of cultivars and synonyms, legal protection, and parentage analysis.
Different cultivars of Vitis vinifera vary in their potential to form embryogenic tissues. The WUSCHEL (WUS)-related homeobox (WOX) genes have been shown to play an important role in coordinating the gene transcription involved in the early phases of embryogenesis. The expression dynamics of 12 VvWOX genes present in the V. vinifera genome in embryogenic and other tissues of 'Chardonnay' were analysed. In order to understand the influence of WOX genes on the somatic embryogenic process, their expression profiles were compared in two cultivars of V. vinifera ('Chardonnay' and 'Cabernet Sauvignon') that show different aptitudes for embryogenesis. The expression of all VvWOX genes was influenced by culture conditions. VvWOX2 and VvWOX9 were the principal WOX genes expressed during the somatic embryogenesis process, and the low aptitude for embryogenesis of 'Cabernet Sauvignon' was generally correlated with the low expression levels of these VvWOX genes. VvWOX3 and VvWOX11 were strongly activated in correspondence to torpedo and cotyledonary stages of somatic embryos, with low expression in the earlier developmental stages (pre-embryogenic masses and globular embryos) and during embryo germination. VvWOX genes appeared to be key regulators of somatic embryogenesis in grapevine, and the regulation of these genes during early phases of somatic embryogenesis differed between the two cultivars of the same species.
In this work, 18 microsatellite loci were developed in the European hazelnut (Corylus avellana L.) using three enriched genomic libraries. They were evaluated on a set of 20 accessions of this species on the basis of number of alleles (mean: 7.1), expected heterozygosity (mean: 0.67), power of discrimination (mean: 0.77) and polymorphism information content (mean: 0.64). Cross‐species transferability was evaluated using seven other Corylus species. All primer pairs amplified in all species, except for CaT‐C505 in Corylus ferox and CaT‐A114 in Corylus californica.
MicroRNAs (miRNAs) are small non-coding RNAs that regulate the post-transcriptional control of several pathway intermediates, thus playing pivotal roles in plant growth, development and response to biotic and abiotic stresses. In recent years, the grapevine genome release, small(s)-RNAseq and degradome-RNAseq together has allowed the discovery and characterisation of many miRNA species, thus rendering the discovery of additional miRNAs difficult and uncertain. Taking advantage of the miRNA responsiveness to stresses and the availability of virus-free Vitis vinifera plants and those infected only by a latent virus, we have analysed grapevines subjected to drought in greenhouse conditions. The sRNA-seq and other sequence-specific molecular analyses have allowed us to characterise conserved miRNA expression profiles in association with specific eco-physiological parameters. In addition, we here report 12 novel grapevine-specific miRNA candidates and describe their expression profile. We show that latent viral infection can influence the miRNA profiles of V. vinifera in response to drought. Moreover, study of eco-physiological parameters showed that photosynthetic rate, stomatal conductance and hydraulic resistance to water transport were significantly influenced by drought and viral infection. Although no unequivocal cause–effect explanation could be attributed to each miRNA target, their contribution to the drought response is discussed.
Grapevine rupestris stem pitting-associated virus (GRSPaV) is a widespread virus infecting Vitis spp. Although it has established a compatible viral interaction in Vitis vinifera without the development of phenotypic alterations, it can occur as distinct variants that show different symptoms in diverse Vitis species. The changes induced by GRSPaV in V. vinifera cv 'Bosco', an Italian white grape variety, were investigated by combining agronomic, physiological, and molecular approaches, in order to provide comprehensive information about the global effects of GRSPaV. In two years, this virus caused a moderate decrease in physiological efficiency, yield performance, and sugar content in berries associated with several transcriptomic alterations. Transcript profiles were analysed by a microarray technique in petiole, leaf, and berry samples collected at véraison and by real-time RT-PCR in a time course carried out at five grapevine developmental stages. Global gene expression analyses showed that transcriptomic changes were highly variable among the different organs and the different phenological phases. GRSPaV triggers some unique responses in the grapevine at véraison, never reported before for other plant-virus interactions. These responses include an increase in transcripts involved in photosynthesis and CO(2) fixation, a moderate reduction in the photosynthesis rate and some defence mechanisms, and an overlap with responses to water and salinity stresses. It is hypothesized that the long co-existence of grapevine and GRSPaV has resulted in the evolution of a form of mutual adaptation between the virus and its host. This study contributes to elucidating alternative mechanisms used by infected plants to contend with viruses.
‘Nebbiolo’ (Vitis vinifera) is among the most ancient and prestigious wine grape varieties characterised by a wide genetic variability exhibited by a high number of clones (vegetatively propagated lines of selected mother plants). However, limited information is available for this cultivar at the molecular and genomic levels. The whole-genomes of three ‘Nebbiolo’ clones (CVT 71, CVT 185 and CVT 423) were re-sequenced and a de novo transcriptome assembly was produced. Important remarks about the genetic peculiarities of ‘Nebbiolo’ and its intra-varietal variability useful for clonal identification were reported. In particular, several varietal transcripts identified for the first time in ‘Nebbiolo’ were disease resistance genes and single-nucleotide variants (SNVs) identified in ‘Nebbiolo’, but not in other cultivars, were associated with genes involved in the stress response. Ten newly discovered SNVs were successfully employed to identify some periclinal chimeras and to classify 98 ‘Nebbiolo’ clones in seven main genotypes, which resulted to be linked to the geographical origin of accessions. In addition, for the first time it was possible to discriminate some ‘Nebbiolo’ clones from the others.
Hazelnut (Corylus avellana L.) is a traditional nut crop in southern Europe. Germplasm exploration conducted on-farm in five countries (Portugal, Spain, Italy, Slovenia, and Greece) identified 77 landraces. The present work describes phenotypic variation in nut and husk traits and investigates genetic relationships using ten simple sequence repeat (SSR) markers among these landraces, 57 well-known references cultivars, and 19 wild accessions. Among the 77 landraces, 42 had unique fingerprints while 35 showed a SSR profile identical to a known cultivar. Among the 42 unique landraces, morphological observations revealed high phenotypic diversity, and some had characteristics appreciated by\ud the market such as nut round and caliber. Analysis of genetic relationships and population structure allowed investigation of the origin and spread of the cultivated germplasm in southern Europe. Our results indicate the existence of three primary centers of diversity in the Mediterranean basin: northwestern Spain (Tarragona) and southern Italy (Campania) in the West and Black Sea (Turkey) in the East. Moreover, the data suggest the existence of secondary gene pools in the Iberian (Asturias) and Italian (Liguria and Latium) Peninsulas, where\ud local varieties were recently domesticated from wild forms and/or from introduced ancient domesticated varieties
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