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…

Shift of Nitrogen Resources by biotic interaction in grapevine

Grape phylloxera (Daktulosphaira vitifoliae Fitch), a monophagous pest of the grapevine, induces nodosities on the roots through its sap-sucking activity.

Read More

Field performance of red and white “pilzwiderstandsfähige” (PIWI) cultivars in the south of Uruguay

As knowledge about the oenological potential of disease-tolerant grape varieties (PIWI) continues to grow and consumer demand for product safety and sustainable production increases, more governments worldwide are permitting the cultivation of these varieties [1].

Read More

Enological characters of thirty vines in four different zones of Tuscany

In the last few years the development of HPLC techniques together with multivariate statistical methods allowed to set methodics of large discriminant and classing efficacy in the study of wine-grapes.

Read More

Dry leaf hyperspectral reflectance predicts leaf elemental composition in grafted hybrids

Elemental composition, measured as the concentrations of different elements present in a given tissue at a given time point, is a key indicator of vine health and development. While elemental composition and other high-throughput phenotyping approaches yield tremendous insight into the growth, physiology, and health of vines, costs and labor associated with repeated measures over time can be cost-prohibitive. Recent advances in handheld sensors that measure hyperspectral reflectance patterns of leaf tissue may serve as an affordable proxy for other types of phenotypic data, including elemental composition. Here, we ask if reflectance patterns of dried Chambourcin leaf tissue from an experimental grafting vineyard can predict the known elemental composition of those leaves.

Read More

Impact of crop load management on terpene content in gewürztraminer grapes

Context and purpose of the study ‐ Crop load management by cluster thinning can improve ripening and the concentration of key metabolites for grape and wine quality. However, little work has been done on testing the impact of crop load management on terpene content of white grapes. The goal of the study was to assess if by reducing crop load via cluster thinning growers can increase terpene concentration of grapes, as well as to test if the timing of thinning application affects terpene concentration.

Read More