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Projects / Programmes source: ARIS

CRISPR/CAS9-mediated targeted mutagenesis for resistance of grapevine and potato against phytoplasmas

Research activity

Code Science Field Subfield
4.03.01  Biotechnical sciences  Plant production  Agricultural plants 

Code Science Field
4.01  Agricultural and Veterinary Sciences  Agriculture, Forestry and Fisheries 
Keywords
Candidatus Phytoplasms solani, CRISP/cas9, DMR6, grapevine, modelling, phytoplasma, potato, resistance, susceptibility genes
Evaluation (rules)
source: COBISS
Points
7,499.29
A''
1,384.66
A'
3,312.35
A1/2
4,281.9
CI10
10,508
CImax
261
h10
50
A1
26.01
A3
13.79
Data for the last 5 years (citations for the last 10 years) on April 19, 2024; A3 for period 2018-2022
Data for ARIS tenders ( 04.04.2019 – Programme tender, archive )
Database Linked records Citations Pure citations Average pure citations
WoS  499  11,962  10,257  20.56 
Scopus  635  15,946  13,644  21.49 
Researchers (16)
no. Code Name and surname Research area Role Period No. of publicationsNo. of publications
1.  36320  PhD Špela Alič  Biology  Researcher  2020 - 2021  82 
2.  34130  PhD Anna Coll Rius  Biochemistry and molecular biology  Researcher  2020 - 2024  163 
3.  08280  PhD Marina Dermastia  Biology  Head  2020 - 2024  872 
4.  23611  PhD Tanja Dreo  Biotechnology  Researcher  2020 - 2024  507 
5.  12688  PhD Kristina Gruden  Biotechnology  Researcher  2020 - 2024  985 
6.  08949  PhD Nada Lavrač  Computer science and informatics  Researcher  2020 - 2024  867 
7.  36323  PhD Tjaša Lukan  Biology  Researcher  2020 - 2021  125 
8.  53799  Martin Marzidovšek  Computer science and informatics  Researcher  2022 - 2024  34 
9.  36912  PhD Dragana Miljković  Computer science and informatics  Researcher  2020  71 
10.  29539  PhD Vid Podpečan  Computer science and informatics  Researcher  2020 - 2024  103 
11.  31844  PhD Senja Pollak  Linguistics  Researcher  2020 - 2024  288 
12.  18467  PhD Maruša Pompe Novak  Biotechnology  Researcher  2020 - 2024  291 
13.  55750  Rebeka Strah  Biology  Researcher  2021 - 2024  14 
14.  52066  PhD Blaž Škrlj  Computer science and informatics  Researcher  2020 - 2024  126 
15.  50566  PhD Špela Tomaž  Biotechnology  Researcher  2020 - 2021  28 
16.  27522  PhD Anže Županič  Systems and cybernetics  Researcher  2020 - 2024  179 
Organisations (2)
no. Code Research organisation City Registration number No. of publicationsNo. of publications
1.  0105  National Institute of Biology  Ljubljana  5055784  13,256 
2.  0106  Jožef Stefan Institute  Ljubljana  5051606000  90,682 
Abstract
Grapevine (Vitis vinifera L.) is an important crop worldwide and production of grapes in 2017 was more tahn 74 million tons, from which approximately one third were produced in Europe. This ranks grape production seventh in agricultural production terms in Europe (http://www.fao.org). This important economic sector is threatened by several grapevine diseases caused by viruses, bacteria, oomycetes, fungi and insects. These pathogens affect production in pre- and post-harvest periods, during processing and export, and they impair the quality of produce. Among the most widespread and problematic grapevine diseases in Europe are flavescence dorée caused by phytopathogenic bacterium FD phytoplasma and bois noir caused by phytoplasma ‘Candidatus Phytoplasma solani’. ‘Ca. P. solani’ also infects plants from the Solanacaeae family and is already a severe threat to production of potato and tomato in neighboring countries, including Austria. In South Africa, the main phytoplasma disease of grapevine, with the same symptoms as developed during bois noir and flavescence dorée, is caused by 'Ca. P. asteris'. Evidence for resistance against phytoplasmas has not been reported in grapevine or potato, but differences in susceptibility between cultivars have been observed, based on disease incidence, symptom severity, as well as plants’ ability to recover. As there is no effective treatment against phytoplasma-infected grapevine or potato available at the moment, obtaining resistant cultivars is crucial. New strategies of plant protection against pathogens include the engineering of loss of function of a plant susceptibility gene (S). Plant genes S are involved in compatible interactions and are required by the pathogen for the infection. Therefore, their deactivation can limit the ability of the pathogen to cause disease. Our transcriptome analysis in phytoplasma-infected grapevine indicates a possible negative role of the DMR6 (downy mildew resistance 6) gene in plant defense. DMR6 encodes oxidoreductase (2-oxoglutarate (2OG)-Fe(II) oxygenase) and functions as a salicylic acid 5-hydroxylase (S5H). It seems a promising S gene candidate target for mutagenesis with a good perspective that its deactivation may increase the resistance to phytoplasmas and also to other pathogens. The main goal of this project is to engineer an improved phytoplasma resistance in grapevine and potato after knockout of the DMR6 gene by the CRISPR/Cas9 system. In parallel, transcriptomic data on phytoplasma infected grapevine, collected in previous studies in Slovenia, Austria and South Africa, will be combined by computing and modelling in order to identify some new suitable candidate susceptibility genes S, other than DMR6, to be targeted for increased resistance of grapevine to phytoplasmas. We suggest an adaptation of DNA-free genetically edited protoplasts, using a direct delivery of purified CRISPR/Cas9 ribonucleoproteins targeting the chosen plant S gene. The regenerated plants will be tested for the effects on phenotype and overall gene expression and compared with the wild type plants. The proposed approach of mutagenesis has better chances that the plants engineered by it will be accepted at the global level. As an alternative procedures we will introduce a nanoparticle-based delivery of CRISPR/Cas9 genetic elements into plant tissues and/or Agrobacterium-mediated transformed S gene knockout plants with the application of the in-house protocols. All regenerated grapevine and potato plants will be inoculated with studied phytoplasmas to test for possible acquired resistance/tolerance against them. Confirmation of an association of the target S gene with an increase in resistance/tolerance toward phytoplasmas will open a plethora of new possibilities in research in the important field of plant immunity, with the potential to lead to novel improved potato and grapevine cultivars.
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