terclim by ICS banner
IVES 9 IVES Conference Series 9 DNA-Free genome editing confers disease resistance in grapevine

DNA-Free genome editing confers disease resistance in grapevine

Abstract

Grapevine (Vitis Vinifera L.), one of the most important cultivated fruit crops, is facing significant challenges due to climate change. Specifically, increasing temperatures negatively impact the physiological traits and disrupt plant phenology. Additionally, increased virulence in pathogen attacks and pests leads to significant yield loss, requiring widespread application of plant protection products. Traditional agronomic practices offer only partial mitigation, requiring the development of precise and effective intervention strategies. The economic worth of viticulture has prompted continuous efforts in grapevine genetic improvement programs, traditionally involving conventional breeding and clonal selection that, however, are complex and time-consuming approaches. Instead, the advent of New Breeding Techniques, especially genome editing via the CRISPR/Cas9 system, presents a promising avenue for the development of tools suitable to mitigate the current viticulture challenges, including fungal diseases. We report the application of a DNA-free genome editing approach to induce targeted mutations in the VviMLO17, a gene associated with powdery mildew susceptibility in grapevine. CRISPR/Cas9 ribonucleoparticles were introduced into protoplasts isolated from embryogenic calli. Through protoplast regeneration, a homozygous edited grapevine plant mutated in the VviMLO17 gene was obtained. This mutation confers resistance to Erysiphe necator, as evidenced by phenotypic analyses that demonstrated a reduced susceptibility to pathogen attack. The success of DNA-free CRISPR/Cas9 application for the improvement of target traits establishes a foundation for promoting viticulture sustainability yet preserving the identity of the grapevine cultivars. This advancement aligns with market and legislative demands, paving the way for a resilient and environmentally conscious winegrowing system.

DOI:

Publication date: June 13, 2024

Issue: Open GPB 2024

Type: Article

Authors

Marianna Fasoli1, Edoardo Bertini2, Erica D’Incà2, Luca Cattaneo1, Stefania Zattoni1, Sara Lissandrini1, Clarissa Ciffolillo1, Annalisa Polverari1, Giovanni Battista Tornielli1,3, Sara Zenoni1*

1 Department of Biotechnology, University of Verona, Verona, Italy
2 EdiVite S.r.l., San Pietro Viminario, Padua, Italy
3 Current address: Department of Agronomy, Food, Natural Resources, Animals and the Environment., University of Padua, Padua, Italy

Contact the author*

Keywords

genome editing DNA-free, CRISPR/Cas9 system, protoplast regeneration, powdery mildew resistance, sustainable viticulture

Tags

IVES Conference Series | Open GPB | Open GPB 2024

Citation

Related articles…

Potential application of indigenous Pichia kluyveri for enhanced wine aroma quality

Aims: In previous work, five indigenous Pichia kluyveri strains, GS1-1, FS-2-7, HS-2-1, C730 and C732, were isolated and selected from spontaneous fermented wines from Ningxia and Gansu. The aims of this study were to 1) evaluate resistance of these strains to environmental stressors that may restrict their growth and the progress of alcoholic fermentation; 2) Investigate their fermentation dynamics; 3) Characterise aroma profiles of Cabernet Sauvignon wines made from mixed cultures of P. kluyveri and Saccharomyces cerevisiae.

Exploring the presence of oligopeptides in wines into identify possible compounds with umami or kokumi properties

Umami is defined as a pleasant and savory taste derived from glutamate, inosinate and guanylate, which are naturally present in meat, fish, vegetables and dairy products. The term “kokumi” refers to a complex flavour sensation, characterized by thickness, fullness and continuity.

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

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.

Integrated approaches for the functional characterization of miRNAs in grapevine

Micro(mi)RNAs are small non-coding RNAs that regulate several pathways and are widely recognised as key players in plant development, tissue differentiation, and many other important physiological processes, including plant adaptation to biotic and abiotic stresses. The release of plant genomes and the application of high throughput sequencing have considerably extended miRNA discovery across many species, including grapevine (Vitis spp.). Despite their relevance in plant development, functional studies in grapevine to clarify the function of miRNAs are not yet available. Through the grapevine genetic improvement platform IMPROVIT at CNR-IPSP (http://www.ipsp.cnr.it/en/thematics/turin-headquarter-thematics/improvit/), we developed integrated approaches to discover miRNA function in grapevine.

WINE SWIRLING: A FIRST STEP TOWARDS THE UNLOCKING OF THE WINE’STASTER GESTURE

Right after the pouring of wine in a glass, a myriad of volatile organic compounds, including ethanol, overwhelm the glass headspace, thus causing the so-called wine’s bouquet [1]. Otherwise, it is worth noting that during wine tasting, most people automatically swirl their glass to enhance the release of aromas in the glass headspace [1]. About a decade ago, Swiss researchers revealed the complex fluid mechanics underlying wine swirling [2]. However, despite mechanically repeated throughout wine tasting, the consequences of glass swirling on the chemical space found in the headspace of wine glasses are still barely known.