CRISPR-Cas9 gene-editing is one of the most powerful tools available to modern science, but genetically-modified organisms (GMOs) in food are subject to some tight regulations. Now, researchers at North Carolina State University have created a new version of CRISPR that lets scientists edit crops without introducing new DNA, meaning they technically aren’t GMOs.
CRISPR-Cas9 allows for precise cut-n-paste edits to DNA in living cells. An RNA guide sequence directs the system to the target section of the genome. Once there, an enzyme, usually Cas9, snips out the sequence then deletes it or replaces it with something else. In this way, scientists can cut out problem genes, such as those that cause disease, or add new beneficial ones, such as giving crops better pest resistance.
For the new study, the researchers tweaked the process to make a “cleaner” edit in plants. It uses a process known as lipofection, where positively-charged lipids are used to build a kind of bubble around the Cas9 and RNA mechanisms. When injected into the organism, this bubble binds to and fuses with the cellular membrane, which pushes the CRISPR system into the cell itself. The method also uses a Cas9 protein itself, rather than the Cas9 DNA sequence.
The team tested the method by introducing fluorescent proteins into tobacco plants. And sure enough, after 48 hours the edited plants were glowing, indicating it had worked.
The new method has a few advantages over existing ones, the team says. It’s easier to target the desired genetic sequence, and opens up new crops that couldn’t be edited with existing methods. Plus, the protein only lasts for a few days before degrading, which reduces off-target edits.
But the most important advantage is that the resulting crops aren’t considered GMOs. Since the new method doesn’t use Cas9 DNA, it doesn’t introduce foreign DNA into the plant, which is an important distinction.
“This was the first time anyone has come up with a method to deliver the Cas9 protein through lipofection into plant cells,” says Wusheng Liu, lead author of the study. “Our major achievement was to make that happen. Also, since many consumers prefer non-GMO specialty crops, this method delivers the Cas9 protein in a non-GMO manner.”
GMO or non-GMO?
As useful as genetic engineering can be, the term “GMO” has negative connotations for many people, who believe there are health concerns with eating these crops or meats. Other problems include the chance of modified plants or animals escaping into the wild, where they can spread their new genes to the native population, affecting ecosystems.
As such, the US Department of Agriculture (USDA) and the Food and Drug Administration (FDA) have regulations on which edited crops and animals are allowed in food. And they’ve decided that the line is drawn at introducing foreign genes into an organism.
It makes sense. Humans have been genetically-engineering plants and animals for millennia, through selective breeding. Many of our most widely-eaten crops are bigger, tastier, and easier to eat or grow, to the point that they hardly resemble their wild counterparts anymore.
CRISPR and other gene-editing tools can be the next generation of this process. By removing problematic genes or ensuring that specific ones are turned on or off, scientists aren’t really creating anything new. Some individuals naturally have mutations that do the same thing – all the scientists are really doing is removing the element of chance, genetically.
In 2015, a new type of salmon became the first genetically engineered animal approved by the FDA for human consumption. In 2016, a Swedish scientist grew, harvested and served up CRISPR cabbage after approval by the Swedish Board of Agriculture. In both cases, the products were allowed because they were functionally identical to wild-type organisms – the scientists had just chosen beneficial genes from an existing natural pool, without introducing foreign DNA.
That said, the rules aren't the same everywhere. In 2018 the Court of Justice of the European Union somewhat controversially ruled that tough GMO laws applied to crops that had been edited even if new DNA hadn't been inserted. The issue will likely remain fragmented, but for the NC State team at least, their crops aren't GMOs according to their own country's regulations.
Alternatives
However, there are still some hurdles to overcome before the new method becomes viable. The team says that lipofection can only be done if the outer wall of the plant cell is removed first. This kind of plant cell, known as a protoplast, allows scientists to more easily tweak the genes, but it isn’t possible in all types of crops, and even when it does work, it’s a complex process.
Instead, the researchers are exploring other options that don’t require removing the cell wall at all. One such alternative is to use CRISPR to introduce the Cas9 protein into pollen grains, which can then go on to fertilize another plant. Some of the offspring will have the required genetic edits from day one.
The researchers plan to investigate this latter method in tomatoes and hemp first, before moving onto others.
The new study was published in the journal Plant Cell Reports.
Source: NC State University
