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…

Unveiling the viticultural heritage of Rachaya, Lebanon: genetic, ampelographic and chemical profiling of local grape varieties

Located in the western bekaa region of lebanon, rachaya is known for its extensive history of viticulture and ideal mediterranean conditions for grapevine cultivation. Despite the area’s importance in grapevine production, there is limited knowledge about the local grape varieties grown there. In this work, we conducted a detailed analysis of five local grapevine varieties in the area, obeidy, kassoufi, foddeh, aswadi, and maryami, of which little is currently known, in comparison to a set of international varieties.

Read More

Health space in vine spa in the world

This elaboration presents vine spa has precious contribution of social development health and well being in culture of wine regions. The majority of the vine-spas in the world draw raw materials from the vineyard; both for cosmetics treatments and for dishes in their restaurants. Vitis vinifera vine provides fresh grapes for dishes and massages, seeds and oil from the seeds, as well as the leaves, and its extracts, and above all the wine.

Read More

Investigating the carbon sequestration potential in vineyard soils–the SUSTAIN project

The SUSTAIN project aims at assessing the soil organic carbon (SOC) stock and vulnerability in vineyard in a climate change scenario.

Read More

Entomopathogenic nematodes application for controlling Lobesia botrana in grapevine and their impact on grapevine quality 

Entomopathogenic nematodes (EPN) are well-known biological control agents combined with specific adjuvants that now allow their use against aerial pests. Lobesia botrana (Lepidoptera: Tortricidae) is one of the major harmful pests detected in worldwide vineyards. Previous studies demonstrated that the EPNs Steinernema feltiae and S. carpocapsae could control L. botrana. The hypothesis was that the best combination of EPN-adjuvant/timing (season/temperatures) will support the use of EPN in the vineyard against L. botrana with no impact on the grape performance.

Read More

Innovative red winemaking strategy: biosurfactant-assisted extraction and stabilization of phenolic compounds

The color is the first attribute perceived by consumers and a major factor determining the quality of red wines. This depends mainly on the content of grape anthocyanins and their extraction into the juice/wine during winemaking. Furthermore, these compounds can undergo reactions that influence the chemical and sensory characteristics of the wine. Monomeric forms are prone to oxidation and adsorption on solid parts.

Read More