Preliminary steps of a protocol to isolate transcription factors bound to a specific DNA locus in grapevine using CRISPR-dCas9 system

Pilzwiderstandsfähige (PIWI) are disease resistant Vitis vinifera interspecific hybrid varieties that are receiving increasing attention for ability to ripen in cool climates and their resistance to grapevine fungal diseases. Wines produced from these varieties have not been characterized, especially regarding their macromolecular composition. This study characterised and quantified colloid-forming molecules (proteins, polysaccharides and phenolics) of red PIWI wines produced in the UK. METHODS: In 2019 6 wines were made from the PIWI varieties Rondo, Cabernet Jura, Cabernet Cortis, Cabernet Noir, Regent and Cabertin grown at the Plumpton Rock Lodge Vineyard in Sussex (UK) and harvested at similar level of maturity (TSS, pH and TA). All juice was chaptalized to the same potential alcohol of 12%. Small scale winemaking (1L) was performed in quadruplicate using Bodum® coffee plungers to manage maceration [1]. Residual sugar content, pH, and titratable acidity were monitored during fermentation. For finished wines, the protein and polysaccharide content was measured by HPLC-SEC [2], while the total phenolic content was assessed using the Folin-Ciocalteau method [3]. The protein profile of the wines was further investigated by SDS-PAGE [4]. RESULTS: Fermentations (n=24) were all carried out to completion within 8 days.

Measurements of parameters spatialy positionned, with on line sensors mounted on classical machinery or airborne imagery is no more a problem in viticulture. In a short time, high resolution data dedicated to the assessment of the vine characteristics, the soil, the harvest, etc. will become a reality.

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Grapevine cultivars can be unequivocally typed by both physical differences (ampelography) and genetic tests. However due to their very similar characteristics, the identification of clones within a cultivar relies on the accurate tracing of supply records to the point of origin. Such records are not always available or reliable, particularly for older accessions. Whole genome sequencing (WGS) provides the most highly detailed methodology for defining grapevine cultivars and more importantly, this can be extended to differentiating clones within those cultivars.

The Champagne vineyard extends over 35,300 ha under the Appellation d’Origine Contrôlée, of which 30,000 are in production. It mainly covers 3 departments: in order of importance, Marne (68% of the appellation area), Aube (22%) and Aisne (10%), and more anecdotally Haute Marne and Seine and Mame. It is a young vineyard (for more than half of the surface, the winegrowers have the experience of only one generation of vines), and fragmented (more than half of the exploitations extend over less than 1 ha; the average size of a cadastral parcel is 12 ares).