terclim by ICS banner
IVES 9 IVES Conference Series 9 Open-GPB 9 Open-GPB-2024 9 Flash - New biotechnological tools 9 Enhancing hydric stress tolerance by editing the VviMYB60 promoter with CRISPR/Cas9 

Enhancing hydric stress tolerance by editing the VviMYB60 promoter with CRISPR/Cas9 

Abstract

Climate change presents increasing challenges to viticulture, particularly with rising water stress contributing significantly to yield losses and damages. The identification of the MYB60 transcription factor, which regulates stomatal opening and closing in Arabidopsis thaliana and Vitis vinifera, offers potential solutions. Notably, knockout studies in Arabidopsis have shown reduced stomatal opening and increased drought tolerance in myb60 mutants. Additionally, the grapevine ortholog, VviMYB60, can restore the wild-type phenotype of Arabidopsis myb60 mutants. Further investigation of the Arabidopsis promoter region has revealed that mutations in DOF motifs lead to reduced expression of AtMYB60.

Utilizing the advancements in CRISPR/Cas9 genome editing, this project aims to modify the VviMYB60 promoter region to lower gene expression, thereby reducing stomatal opening in grapevines. Binary vectors for genome editing were constructed to target two specific regions of the VviMYB60 promoter. Agrobacterium-mediated transformation was performed on Chardonnay embryogenic calli, resulting in the successful regeneration of plants under selection conditions. Sanger sequencing analysis of the targeted region confirmed the occurrence of genetic edits in four of the six lines analyzed so far.

To further characterize the edited lines, next-generation sequencing will be utilized, providing a more comprehensive understanding of the mutations, as well as gene expression of VviMYB60 will be evaluated to confirm that the editing reduces its expression. Morphological and physiological parameters will be measured after acclimatation in greenhouse and finally these edited lines will undergo drought tolerance testing to assess their performance.

DOI:

Publication date: June 13, 2024

Issue: Open GPB 2024

Type: Article

Authors

Manuela Campa1*, Loredana Moffa2, Luca Nerva2, Walter Chitarra2, Johan Burger1

1 Genetics Department, Faculty of Agrisciences, Stellenbosch University, 7600 Stellenbosch, South Africa
2 Research Centre for Viticulture and Enology, Council for Agricultural Research and Economics (CREA-VE), Via XXVIII Aprile 26, 31015 Conegliano (Italy)

Contact the author*

Keywords

CRISPR/Cas9, VviMYB60, promoter, drought tolerance, stomatal regulation

Tags

IVES Conference Series | Open GPB | Open GPB 2024

Citation

Related articles…

A comprehensive and accurate annotation for the grapevine T2T genome 

Addressing the opportunities and challenges of genomics methods in grapevine (Vitis vinifera L.) requires the development of a comprehensive and accurate reference genome and annotation. We aimed to create a new gene annotation for the PN40024 grapevine reference genome by integrating the highly accurate and complete T2T assembly and the manually curated PN40024.v4 annotation. Here, we present a novel workflow to enhance the annotation of the T2T genome by incorporating past community input found in PN40024.v4. The pipeline’s containerization will improve the workflow’s reproducibility and flexibility, facilitating its inclusion as a shared workflow on the Grapedia portal, the grapevine genomics encyclopedia.

Mapping aromatic profiles of Chardonnay and Sangiovese wines in grafting combination with new rootstocks

Rootstocks play a key role in the adaptation of grapevine to environmental conditions, affecting phenology, vigour, yield and grape quality.

Chemical and biochemical formation of polysulfides in synthetic and real wines using UHPLC-HRMS

ulfur compounds in wine have been studied for several years due to their impact on wine flavour, but the role of polysulfides is a recent topic. Polysulfides in wine are formed when two sulfhydryl groups oxidize, especially in presence of elemental sulfur or metal catalysts from field treatment residues (Ugliano et al. 2011). These compounds are odourless, but can degrade during storage and affect the wine quality. The mechanism of their formation is still largely unknown but different chemical and biochemical pathways have been suggested. Disulfides from cysteine (Cys) and glutathione (GSH) have been revealed in model wines (Kreitman et al. 2016) and more recently also higher polymerized forms in real wines (Van Leeuwen et al. 2020). Volatile varietal thiols like 3-mercaptohexanol (3MH) and 4-mercaptopentanone (4MMP) – flavour compounds with tropical or fruity notes – could undergo similar reactions, also with Cys and GSH, subsequently losing their flavour property (fate). Even more concerning is the possible release of H2S from polysulfides during storage, leading to undesired off-flavours (Sarrazin et al. 2010).

Caratterizzazione delle produzioni vitivinicole dell’ area del Barolo: un’esperienza pluridisciplinare triennale (5)

[lwp_divi_breadcrumbs home_text="IVES" use_before_icon="on" before_icon="||divi||400" module_id="publication-ariane" _builder_version="4.19.4" _module_preset="default" module_text_align="center" module_font_size="16px" text_orientation="center"...

A lower rate of grape berry transpiration delays ripening and reduces flavonoid content

Exposing berries to solar radiation improves most berry composition traits. Many of these effects have been linked to photomorphogenic mechanisms and berry temperature.