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

Wine is the result of the alcoholic fermentation (AF) of grape must. Besides AF, wine can also undergo the malolactic fermentation (MLF) driven out by lactic acid bacteria (LAB). Among LAB, Oenococcus oeni and Lactiplantibacillus plantarum are the dominant species in wine. Even if O. oeni is the most common LAB undergoing MLF in wine, due to its high tolerance to wine conditions, L. plantarum can be used to undergo MLF in must. The moderate tolerance of L. plantarum to low pH and ethanol, may compromise the fermentative process in harsh wines.

In Tuscany region the Rufina is a district of Chianti D.O.C.G. positioned in Val di Sieve, 20 km north east from Florence.

The Pomerol vineyard, located 35 km east of Bordeaux, covers around 800 ha on the left bank of the Isle. There is a system of fluvial terraces with more or less coarse gravel and pebble spreading, resting on a Tertiary substratum ranging from the Middle to Upper Eocene to the Lower Oligocene (Dubreuilh, 1993). This interweaving of terraces of varying thickness results in a brutal superposition of differentiated materials which give rise to various types of soil. Several site studies in this sector of the Libounais show significant morphological and analytical differences from one point to another (Guilloux et al ., 1978; Duteau, 1982; Van Leeuwen et al.., 1989). The distribution of the soils of the Pomerol vineyard was studied and resulted in a cartography at 1/25000th (Merouge, 1995).

Climate change is causing a gap between the technological and phenolic maturity of grapes, resulting in wines with high alcohol content and low polyphenol concentration. Another phenomenon associated with high temperatures and whose effect is more pronounced if the harvest is delayed is the decrease in the acidity of the grapes, mainly in malic acid, and an increase in pH caused by the accumulation of potassium derived from the increase in temperature. Therefore, climate change and the effects it causes on the vine leads to unbalanced wines, with high alcohol content and lack of color, with green tannins, astringency and excessively low acidity if not corrected.

The use of cover crops under the vines might be an alternative to the use of herbicides or tillage, improving grapevine quality and soil characteristics. The aim of this research was to study the implications of different management strategies of the soil under the vines (herbicide, cultivation or cover crops) on grapevine growth, water and nutritional status, gas exchange parameters and belowground microbial communities.
The experimental design consisted in 4 treatments applied on 35L-potted Tempranillo vegetative grapevines with 10 replicates each grown in an open-top greenhouse in 2022 and 2023. Treatments included two cover crop species (Trifolium fragiferum and Bromus repens), herbicide (glyphosate al 36%) and an untreated control.