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We have already seen many different uses for gene editing CRISPR – the precise cutting and gluing of particular genes into DNA. The researchers have come up with an improvement in the process that could allow simultaneous editing of tens or even hundreds of genes.
According to the team, this could open up all kinds of new possibilities, allowing scientists to reprogram cells on a larger scale and in a more sophisticated way: when studying complex genetic disorders, for example, or when the attempt to replace damaged cells with healthy cells.
For the most part, CRISPR techniques only alter one gene at a time, although up to seven genes have been edited together. According to this latest study, the new method can reach 25 targets simultaneously in genes.
"Our method allows us, for the first time, to systematically change entire gene networks in one step," says biochemist Randall Platt of ETH Zurich in Switzerland. "With this new tool, we, as well as other scientists, can achieve what we could only dream of in the past."
The key to the new multiple targeting system is a plasmid-stabilized RNA structure, or circular DNA molecule, capable of containing and processing additional RNA molecules. These RNA molecules act as address markers to target gene sites. Therefore, the higher the plasmid can carry, the greater the number of parts of a cell that scientists can target.
In addition to the RNA address molecules, the plasmid carries a Cas enzyme which performs the binding and cleavage work itself. Cas9 is the enzyme used most often, but here scientists have turned to Cas12a, an enzyme that has already been shown to improve the accuracy of CRISPR editing.
In their experiments, the scientists managed to insert their new plasmid into human cells of the laboratory.
These changes to the standard CRISPR process could allow scientists to perform more widespread gene editing. The genes and proteins inside cells interact in an extremely complex way. Sometimes only one cut or change at a time can be limiting.
For example, the new technique could mean that the activity of certain genes can be increased at the same time as the activity of other genes is reduced – and that all these operations can also be planned with one more high accuracy.
There is a trap here, though. We do not know exactly how other changes might occur in the body being modified. As we have seen in the past, there may be unexpected side changes, and the more genes you change, the higher the risk.
"Direct repetitive sequences and spacers containing complementary sequences [..] could generate complex secondary RNA structures affecting CRISPR RNA processing in cells, "writes the team in its article.
"Therefore, complementary regions in pre-CRISPR RNA need to be considered to improve the maturation of CRISPR RNA." Future work to overcome these limitations will open up many applications for the engineering of CRISPR. highly multiplexed genome. "
We have already seen CRISPR used to eliminate the genes responsible for the disease and to eliminate superbugs. Scientists say that a lot of other things are coming and that they now have an even more versatile and comprehensive toolbox.
"Our method provides a powerful platform for studying and orchestrating sophisticated genetic programs underlying complex cell behaviors," the team writes.
The search was published in Nature methods.
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