Genome editing for improved vines
The effects of climate change and population growth open new scenarios for agriculture and set new goals for researchers and farmers. In fact, today’s increase in agricultural production cannot be separated from the reduction of its environmental impact. In this perspective, green biotechnology research is focused on studying plant genomes to improve adaptability and resistance to different environmental conditions and pathogens/parasites, without altering the genetic characteristics of the cultivated varieties. The goal is to grow stronger and more adaptive plants and enable farmers to become less dependent on chemicals to protect crops. The picture just described is the focal point of the NeosVine research project, started by IGA Technology Services, a spin-off of the Institute of Applied Genomics of Udine (IGA) in collaboration with the Vivai Cooperativi Rauscedo, granted by the Italian Autonomous Region Friuli Venezia Giulia.
NeosVine is a research project designed specifically for the grapevine. Grapevine growth phases are very sensitive to climatic and environmental conditions. Water abundance or shortages as well as heat waves can alter the fruit ripening process, compromising the grapes quality. The berries that ripen too early in excessive heat conditions have higher sugar content and a lower level of acidity, influencing the organoleptic characteristics of the final product, i.e. flavor, perfume and color of grapes and wine. In addition, high temperatures associated with heavy rainfall create a warm-humid environment favorable for the proliferation and spread of diseases and pests.
"The spread of pathogens is a problem of primary importance for the future of wine production” explains Eleonora Paparelli, researcher at IGA Technology (https://igatechnology.com/). "For this reason, IGATech has been working for years to obtain fungi-resistant plants, that require a reduced use of pesticides, for the benefit of the environment, farmers and consumers."
An extraordinary development of the genome editing in the gene therapy field has a great potential of being extended to the crop improvement. This revolutionary technique, based on CRISPR/Cas9 system, uses a mechanism naturally present in bacteria to induce targeted mutations by adding or removing just a few nucleotides in one of individual's genes. "No foreign genes are inserted in the plant’s DNA during the process.", explains Paparelli "Instead, a process, which corrects the DNA sequence, is activated". What acts as an immune system in bacteria can be used in non-bacterial cells as a sort of scissors: the Cas9 protein, in fact, is able to cut the DNA in one precise point of the genome. This allows to accurately perform a specific genetic modification, eliminating or replacing sequence that can be harmful or undesired (responsible of a certain biological function in the plant), keeping the rest of the genome intact.
In the case of the grapevine, at present there are only pilot experimental CRISPR/Cas9 protocols, further studies are needed to establish this technique as an easy-to-use tool to produce improved vines. During NeosVine IGATech researchers intend to define a protocol that applies CRISPR/Cas9 for the improvement of grapevine genetic characteristics in a shorter time compared to traditional breeding approaches and at lower costs.
"Setting up the protocol means, at this stage, understand above all how to regenerate the plant after the cut introduced by CRISPR/Cas9 scissor, which is performed on cells similar to embryonic ones”, specifies the researcher. Current research teaches us that once the mutation is induced the main problem is the plant reconstitution. "When it comes to regeneration, the grapevine is a quite recalcitrant species. A process can last even twelve months," explains Paparelli "thus some molecular biology techniques are difficult to apply”. For this reason, researchers chose to edit genes involved in the color development or size of the leaves, characteristics immediately visible in the just germinated seedlings and not phenotypes linked to the fruit which appear in the adult plant. In this way, it will be possible to prove the efficiency of the technique already after one year.
Initially, the researchers will isolate the grapevine cells and apply the CRISPR/Cas9. In a second phase, they will induce the regrowth of the plant from the transformed cells and finally sequence the genome of the regenerated plant to verify the presence of the induced mutation, the position of the modification, the absence of off-target mutations and the impact of the modification on the plant phenotype. "Once the protocol is set up," explains Paparelli, "the research will continue with the editing of pathogen resistance or stress response genes, those of tangible interests for the cultivation". Since the birth of agriculture, the improvement of plants to respond to environmental or anthropogenic factors is the central activity of farmers. Traditional techniques, such as breeding is already used to improve existing grapevines and produce new varieties. The challenge now is to speed-up this process and be less drastic in introducing changes.
--- Fernanda Marchiol