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.

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…

Nuove soluzioni e strumenti per l’agricoltura e la viticoltura di precisione

GEOSPHERA s. r. l. e TERR.A.IN. CNS, forti della grande esperienza dei loro collaboratori nell’ambito delle scienze naturali, della geologia, della geofisica e dell’informatica, garantiscono risposte innovative alle problematiche della moderna agricoltura rivolgendosi direttamente ai viticoltori, ai commercianti vinicoli ed ai liberi professionisti.

Mining belowground and aboveground microbiome data to identify microbial biomarkers of grapevine health and yield

Vineyards are home to a wide diversity of microorganisms that interact with plants and with each other.

Rootstock x environment interaction shapes shoot system phenotypic variation in grafted ‘Chambourcin’

Recent advances in phenomics and transcriptomics have the enhanced capacity for understanding how clonally propagated perennial crops like grapevines respond to their environments seasonally and over the course of multiple years. Because most grapevines are grafted, above-ground grapevine traits reflect scion genotype and its interaction with the local environment. In addition, traits expressed by the scion reflect rootstock genotype and how that rootstock is interacting with its environment seasonally and across years. To investigate rootstock x environment interaction on shoot systems in grafted grapevines we characterized comprehensive phenotypic variation in an experimental vineyard in Mount Vernon, Missouri, USA where the grapevine cultivar ‘Chambourcin’ is growing on its own roots and is grafted to three different rootstocks (‘1103P’, ‘3309C’, ‘SO4’).

Canopy photosynthetic activity and water relations of Syrah/R99 as affected by row orientation on a particular terroir

L’activité photosynthétique et les relations hydriques de plantes de Syrah sur R99 un mois après la véraison ont été étudiées dans un vignoble de la région de Stellenbosch. Le vignoble, planté à 2,75 entre rangs et 1,5 m sur le rang, sur un sol de type Glenrosa, était en pente et exposé a l’ouest: pour les rangs on avait adopté une orientation nord

On the losses of dissolved CO2 from laser-etched champagne glasses under standard tasting conditions

Under standard champagne tasting conditions, the complex interplay between the level of dissolved CO2 found in champagne, its temperature, the glass shape, and the bubbling rate, definitely impacts champagne tasting by modifying the neuro-physico-chemical mechanisms responsible for aroma release and flavor perception. Based on theoretical principles combining heterogeneous bubble nucleation, ascending bubble dynamics and mass transfer equations, a global model is proposed (depending on various parameters of both the wine and the glass itself), which quantitatively provides the progressive losses of dissolved CO2 from laser-etched champagne glasses.