Project description:Background: Grape cultivars and wines are distinguishable by their color, flavor and aroma profiles. Omic analyses (transcripts, proteins and metabolites) are powerful tools for assessing biochemical differences in biological systems. Results: Berry skins of red- (Cabernet Sauvignon, Merlot, Pinot Noir) and white-skinned (Chardonnay, Semillon) wine grapes were harvested near optimum maturity from the same experimental vineyard and Ë?Brix-to-titratable acidity ratio. Identical sample aliquots were analyzed for transcripts by grapevine whole-genome oligonucleotide microarray and RNA-seq technologies, proteins by nano-liquid chromatography-mass spectroscopy, and metabolites by gas chromatography-mass spectroscopy and liquid chromatography-mass spectroscopy. Principal components analysis of each of five Omic technologies showed similar results across cultivars in all Omic datasets. Comparison of the processed data of genes mapped in RNA-seq and microarray data revealed a strong Pearson's correlation (0.80). The exclusion of probesets associated with genes with potential for cross-hybridization on the microarray improved the correlation to 0.93. The overall concordance of protein with transcript data was low with a Pearson's correlation of 0.27 and 0.24 for the RNA-seq and microarray data, respectively. Integration of metabolite with protein and transcript data produced an expected model of phenylpropanoid biosynthesis, which distinguished red from white grapes, yet provided detail of individual cultivar differences. Conclusions: The five Omic technologies were consistent in distinguishing cultivar variation. There was high concordance between transcriptomic technologies, but generally protein abundance did not correlate well with transcript abundance. The integration of multiple high-throughput Omic datasets revealed complex biochemical variation amongst five cultivars of an ancient and economically important crop species. Vitis vinifera L. cv. Cabernet Sauvignon, Chardonnay, Merlot, Pinot Noir, Semillon berries were harvested from Nevada Agricultural Experiment Station Valley Road Vineyard, Reno, NV, USA. Whole-genome microarray analysis was used to assess the transcriptomic response of berry skins at harvest, approximately 24 Â°Brix (2011 vintage). Vines were grown under water deficit and well-watered conditions. At least two clusters harvested from non-adjacent vines were used for each of five experimental replicates.

Project description:Anthocyanins are flavonoid compounds responsible for red/purple colours in the leaves, fruit and flowers of many plant species. They are produced through a multistep pathway which is controlled by MYB transcription factors. VvMYBA1 and VvMYBA2 activate anthocyanin biosynthesis in grapevine (Vitis vinifera) and are non-functional in white grapevine cultivars. In this study, transgenic grapevines with altered VvMYBA gene expression were developed, and transcript analysis was carried out on berries using a microarray technique. The results showed that VvMYBA is a positive regulator of the later stages of anthocyanin synthesis, modification and transport in Shiraz. One up-regulated gene ANTHOCYANIN 3- O-GLUCOSIDE-6â-O-ACYLTRANSFERASE (Vv3AT) encodes a BAHD acyltransferase protein, belonging to a clade separate from most anthocyanin acyltransferases. Functional studies (in planta and in vitro) show that Vv3AT has a broad anthocyanin substrate specificity and can also utilise both aliphatic and aromatic acyl donors, a novel activity for this enzyme family found in nature. In V. vinifera cv. Pinot Noir, a red-berried grapevine mutant lacking acylated anthocyanins, Vv3AT contains a nonsense mutation encoding a truncated protein that lacks two motifs required for BAHD protein activity. Promoter activation assays confirm that Vv3AT transcription is activated by VvMYBA1, which adds to the current understanding of the regulation of the BAHD gene family. The flexibility of Vv3AT to use both classes of acyl donors will be useful in the engineering of anthocyanins in planta or in vitro. Transgenic Chardonnay/Shiraz and non-transformed WT controls were all grown in the same glasshouse in ambient light with a night break. Day and night temperatures were approximately 27oC and 22oC respectively. For microarray experiments, whole berries were sampled from independent transgenic lines: three from transgenic Chardonnay and four from transgenic Shiraz, resulting in three and four biological replicates respectively. Bunches were harvested close to ripeness based on average total soluble solids (TSS, measured as oBrix). This was aimed to be between 20 â 24 oBrix and was determined from TTS of a subsamples from each bunch (Table S53). A sample consisted of all remaining berries from a single bunch except when there were <100 berries in which case more than one bunch was used in the one replicate. Whole berries were immediately frozen in liquid nitrogen. For skin samples, the skins were first removed from fresh berries then frozen in liquid nitrogen. All samples were stored at -80oC.

Project description:Grapevine rupestris stem pitting-associated virus (GRSPaV) is a widespread virus affecting Vitis spp. Although it has established a compatible viral interaction in Vitis vinifera L. without the development of phenotypic alterations, it can occur as distinct variants that show different symptoms in diverse Vitis species. We investigated the changes induced by GRSPaV in V. vinifera cv Bosco, an Italian white grape variety, by combining agronomic, physiological, and molecular approaches, in order to provide comprehensive information about the global effects of GRSPaV. In two consecutive 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 microarray techniques in petiole, leaf, and berry samples collected at véraison and by quantitative real-time RT-PCR (qRT-PCR) in a time course carried out at five relevant 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, such as an increase in transcripts involved in photosynthesis and CO2 fixation, associated with a moderate reduction in the photosynthesis rate and some defence mechanisms, and to an overlap with responses to water and salinity stresses. We hypothesise that the long co-existence between 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. The study was carried out in a vineyard planted in 2002 in Albenga (Liguria), North-West Italy, where a row was established with the white grape cultivar Bosco (V. vinifera L.). Microarray analysis was carried out on leaves, petioles, and berries collected at véraison (E-L35) in 2010. For each of the six GRSPaV-free and six GRSPaV-infected vines selected for the physiological and agronomical parameters evaluation, we collected 6 leaves (3 basal and 3 apical) with the related petioles and 12 berries from 3 different bunches. Samples from each organ were arbitrary pooled in 3 independent biological replicates and total RNA was extracted according to the method described by Gambino et al. (2008).

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Vitis vinifera

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