Thirty years after the marketing of a first genetically modified tomato, resistant to rot, the Flavr Savr, which had disappeared from the market for a long time, super-tomatoes with improved taste and virtues which were obtained by help of new genome editing technologies. These technologies, such as CRISPR-Cas9, which are very different from the transgenesis used in the creation of traditional genetically modified organisms (GMOs), are poised to revolutionize the world of agriculture.
On November 13, Chinese researchers stated in the journal Nature having designed a tomato that is 30% sweeter than commercially grown varieties. To do this, the researchers used CRISPR-Cas9 technology to inactivate two genes present in the tomato genome which control the sugar content of the fruit. By blocking these two genes, the glucose and fructose contents of tomatoes increased by 30%, without affecting fruit size and crop yield.
A tomato rich in gamma-aminobutyric acid (GABA), a compound believed to be able to lower blood pressure and help relax, has also been developed using the CRISPR-Cas9 technique and marketed by the young company Sanatech Seed from the University of Tsukuba, Japan. Sanatech Seed even received the green light in September 2021 to market this red Sicilian tomato containing four to five times more GABA than traditional tomatoes, in this country, where people are fond of foods and drinks enriched with this acid amine. This tomato, in which we have simply neutralized a gene, is the first plant whose genome has been modified by CRISPR-Cas9 to have been commercialized.
“Small adjustment on a gene”
But how do these new genome editing technologies, the main one of which is CRISPR-Cas9, differ from transgenesis used to produce GMOs or, to be more precise, transgenic plants? Essentially, “genome editing techniques make it possible to make a small adjustment, a small adjustment, to a gene that the plant possesses. We do not insert any foreign DNA, unlike what we do for GMOs [traditionnels] “, summarizes Jaswinder Singh. This professor in the Department of Plant Sciences at McGill University gives the example of GMOs, such as BT corn, to which a bacterial gene was added that was not present in the genome of these plants. BT corn contains a gene from the bacteria Bacillus thuringiensiswhich confers resistance to harmful insects, including the moth.
Genome editing only makes “very small changes, such as removing or inserting just a few nucleotides [les unités de base de l’ADN] in a particular gene in the genome of a plant which can contain, like soybean, a billion nucleotides. The genetic background of the plant remains the same. Whereas, to produce a transgenic plant, a new gene is introduced [que l’on appelle transgène] which can contain 10,000 nucleotides, and which, most often, comes from a different species, such as another plant, a fungus, an animal, a bacteria,” specifies François Belzile, genomics researcher at Laval University.
Another fundamental difference between the two approaches is the high precision of genome editing technologies. The main technique, CRISPR-Cas9, is in fact a pair of molecular scissors that is introduced into cells and can be positioned on a particular gene. “We can control the exact location where we send this tool and the Cas9 enzyme only does one thing: cuts the DNA at that precise location. Following this cut, the cell’s repair systems are activated to glue the DNA strands back together. The repair is often not perfectly faithful to what was there before. Some nucleotides will be omitted, added or substituted. We then carry out characterization work to see the effect of these small modifications, which most often will inactivate the function of the targeted gene,” explains Jean-Benoît Charron, professor at the Faculty of Agriculture and Environmental Sciences at the McGill University.
Another way to do this is to provide the CRISPR-Cas9 system with a tiny piece of DNA (called donor DNA) that contains exactly the modification you want to make. This modification will have been identified beforehand during the characterization of the various small mutations induced by CRISPR-Cas9, or following several years of fundamental research. The cell’s repair system then takes care of integrating these few nucleotides into the targeted gene, explains the researcher.
“Genome editing techniques are so precise that once the mutation is made, even though sometimes there may also be other small, non-specific changes [hors de la cible] here and there in the genome, we always remain below the rate of mutations which occurs naturally in cells when they divide and under the effect of certain environmental factors. This is why some governments are ready to authorize them more easily,” adds Mr. Charron.
With these two processes, we can therefore make the desired changes at a defined location in the plant’s genome, whereas when creating a transgenic plant, we do not control where the transgene will be inserted. “It’s completely random. We must then check whether the plant contains the transgene. And if so, where? Because these methods often tended to insert several copies of the transgene, particularly in different locations. The level of precision is therefore significantly lower. All of this could be corrected through selection processes, but it complicated the work,” underlines Mr. Charron.
“Cleaner”
“The beauty of the CRISPR-Cas9 system also lies in the fact that once it has accomplished its task, the cell will degrade it and get rid of it. It is not inserted into the genome of the plant, unlike what happens in the transgenic approach, where there is integration at the genome level. So it’s much cleaner,” he adds.
Genomic editing is carried out in the floral organs, which are in a way the germ cells (ovules and sperm) of the plant, which ensures the transmission of the modification to the next generations.
Finally, the technologies of Today’s sequencing, which allows the complete genome of plants to be sequenced at low cost, gives us the opportunity to remove all doubts about the modifications that were made to plants during the editing of their genome, says Mr. Charron.
They allow us to “document in the finest detail where the changes took place by comparing the variety resulting from genome editing to the initial variety. We can then respond to certain opponents who argue that there are perhaps changes elsewhere than on the target which can prove toxic and prove to them with absolute certainty that out of a billion nucleotides, there are only two nucleotides. less in a specific location, for example,” adds Mr. Belzile.
“All these molecular techniques are just one tool in our box, a very powerful tool in the case of genome editing. But they will never be the solution to everything, they will never become the Holy Grail. We must use them prudently and not neglect all facets of agriculture, whether agricultural practices, the application of herbicides, etc., believes Mr. Charron. The CRISPR-Cas9 system will never replace traditional plant breeding programs that span decades, but it will allow this work to be done much more efficiently and much more quickly. »
