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…

Development of FTIR partial least squares models for polyphenol quantification in red wine during fermentation

Polyphenolic compounds are considered to have a major impact on the quality of red wines. Sensory impact, such as astringency and bitterness, stems directly from tannin composition. Thenceforth, quick analytical measurement of phenolic compounds appears to be a real challenge for winemaking monitoring and process control. Many methods were developed to analyzed polyphenols in wine, but they are time-consuming and require chemistry skills and equipment, not suitable for a rapid routine analysis. A reliable and rapid method to obtain this kind of measurement is Fourier Transform Infrared (FTIR) spectroscopy.

Yield formation and grape composition: more than meets the eye 

Fruit quality in grapes is not well defined but is often depicted as correlating inversely with crop yield. Both fruit yield and composition, however, are made from distinct components that interact in complex ways. Reproductive growth of grapevines extends over two growing seasons. Inflorescences initiated in buds during the previous year differentiate flowers and set and develop berries during the harvest year.

Lactic acid bacteria: A possible aid to the remediation of smoke taint?

With climate change, the occurrence of wildfires has increased in several viticultural regions of the world. Subsequently, smoke taint has become a major issue, threatening the sustainability of the wine industry.

Wine chemical markers assess nitrogen levels in original grape juice

Nitrogen (N) nutrition of the vineyard plays a crucial role in the composition of must and wine, impacting fermentation, as well as the aroma and taste of the final product. N-deficient grape juice can result in increased astringency and bitterness, and a decrease in pleasant aromas in the wine.

TCA – A status report on South African cork closures

Cork taint decreases the commercial value of wine as tainted wines are rejected by consumers. Although other compounds in wine and cork can also be responsible for causing a taint, 2,4,6-trichloroanisole (TCA) is regarded as the primary cause of cork taint. As cork taint is often used in marketing campaigns against natural cork closures,