terclim by ICS banner
IVES 9 IVES Conference Series 9 Optimized protocol for high-quality RNA extraction from grape tissues using sorbitol pre-wash

Optimized protocol for high-quality RNA extraction from grape tissues using sorbitol pre-wash

Abstract

Obtaining high-quality RNA from grape tissues, including berry pulp, berry skins, stems, rachis, or roots, is challenging due to their composition, which includes polysaccharides, phenolic compounds, sugars, and organic acids that can negatively affect RNA extraction. For instance, polyphenols and other secondary metabolites can bind to RNA, making it difficult to extract a pure sample. Additionally, RNA can co-precipitate with polysaccharides, leading to lower extraction yield. Also, sugars and organic acids can interfere with the pH and ionic properties of the extraction buffer. To address these challenges, we optimized a protocol for RNA isolation from grape tissues. Although commercial kits can provide a rapid extraction, they were inefficient for these plant materials. Similarly, protocols that work well for other vegetal tissues were also inefficient and time-consuming on grape tissues. To overcome these limitations, we added a sorbitol pre-wash step to both a three-day long protocol based on LiCl precipitation and a commercial kit. Our results showed that the addition of a sorbitol pre-wash improved multiple parameters: the A260/280 absorbance ratio, integrity and quality (IQ), and RNA integrity number (RIN). Sorbitol played a crucial role in ensuring high-quality RNA extraction from grape tissues. It inhibits RNase, thereby preserving RNA integrity and stability. It also helps in disrupting cellular membranes, facilitating the release of RNA, and maintains the osmotic pressure through hypertonicity, which is beneficial to RNA extraction. By using sorbitol, commercial kits can be used to extract RNA from challenging grape tissues, leading to an efficient and time-saving procedure.

Download the abstract

DOI:

Publication date: June 13, 2024

Issue: Open GPB 2024

Type: Poster

Authors

Annalisa Prencipe1, Antonella Salerno1,2, Marco Vendemia2, Carlo Bergamini2, Margherita D’Amico2, Lucia Rosaria Forleo2, Teodora Basile2, Maria Francesca Cardone2, Antonio Domenico Marsico2, Riccardo Velasco2, Mario Ventura1, Flavia Angela Maria Maggiolini2*

1 Department of Biosciences, Biotechnology and Environment, University of Bari “Aldo Moro”, 70125 Bari, Italy
2 Council for Agricultural Research and Economics – Research Center Viticulture and Enology (CREA-VE), Via Casamassima 148-70010 Turi (Ba), Italy

Contact the author*

Keywords

Vitis vinifera, RNA, sorbitol, extraction protocol

Tags

IVES Conference Series | Open GPB | Open GPB 2024

Citation

Related articles…

Global warming effects on grape growing climate zones within the Rioja Appllation (DOCa Rioja) in north Spain

Aims: The aims of this work were (1) to assess the changes in some of the main bioclimatic indices used for climate viticultural zoning within the Rioja Appellation area in the north of Spain between 1950-2014 (60 years), and (2) to carry out a comprehensive sociological evaluation among grapegrowers and winemakers of this region, to better understand the impact of climate change on their activity, their degree of concern about it and the potential adaptation measures they would be willing to adopt to cope with it in future years.

Read More

Impact of the fumaric acid/glutathione pair addition before bottling on Cabernet Sauvignon wine quality

Over the last decades, climate change and rising temperatures have impacted the wine industry. Wines from warm regions tend to have a higher pH and lower total acidity.

Read More

The use of remote sensing in South-African terroir research

The diversity of soil types in the Western Cape of South Africa leads to high levels of within-vineyard variability. Multispectral remote sensing has received a lot of attention recently in the South-African wine industry in an attempt to identify and deal with this variability.

Read More

Sensory changes in wines associated with the ripening of Grenache grapes from vineyards in different climatic zones

Climate change is introducing a high variability on grape ripening, causing uncertainty, excessive spending on pesticides and eventually frustrating results in terms of the quality of the vintage, with the increasingly frequent appearance of aromatic problems associated with overripeness, raisining and greenness, which sometimes only appear in bottled wines.

Read More

SENSORY PROFILES AND EUROPEAN CONSUMER PREFERENCE RELATED TOAROMA AND PHENOLIC COMPOSITION OF WINES MADE FROM FUNGUSRESISTANT GRAPE VARIETIES (PIWI)

Planting grape varieties with several resistance loci towards powdery and downy mildew reduces the use of fungicides significantly. These fungus resistant or PIWI varieties (acronym of German Pilzwiderstandsfähig) contribute significantly to the 50% pesticide reduction goal, set by the European Green Deal for 2030. However, wine growers hesitate to plant PIWIs as they lack experience in vinification and are uncertain, how consumer accept and buy wines from these yet mostly unknown varieties. Grapes from four white and three red PIWI varieties were vinified in three vintages to obtain four diffe-rent white and red wine styles, respectively plus one rosé.

Read More