Visualisation of genes and proteins interacting. (ETH Zurich / Carlo Cosimo Campa)

In a Dizzying New Upgrade, CRISPR Can Now Edit Several Genes at Once

By David Nield

August 19, 2019

We've already seen many different uses for CRISPR gene editing – the precise cutting and pasting of particular genes in DNA. Now researchers have come up with an improvement to the process that could allow for dozens or even hundreds of genes to be edited simultaneously.

According to the team, this could open up all kinds of new possibilities, enabling scientists to reprogram cells on a larger scale and in more sophisticated ways: when studying complicated genetic disorders, for example, or when attempting to replace damaged cells with healthy ones.

For the most part, CRISPR techniques only modify a single gene at once, though on occasion as many as seven genes have been edited together. According to this latest study, the new method can hit 25 targets within genes simultaneously.

"Our method enables us, for the first time, to systematically modify entire gene networks in a single step," says biochemist Randall Platt, from ETH Zurich in Switzerland. "Thanks to this new tool, we and other scientists can now achieve what we could only dream of doing in the past."

The key to the new multi-targeting system is a stabilised RNA structure in a plasmid, or circular DNA molecule, capable of holding and processing additional RNA molecules. These RNA molecules act as address labels for targeting gene sites, so the more the plasmid can carry, the more parts of a cell scientists can target.

As well as the RNA address molecules, the plasmid carries a Cas enzyme, which does the actual binding and cutting work. Cas9 is the enzyme used most often, but here the scientists switched to Cas12a, an enzyme that's previously been shown to improve the accuracy of CRISPR editing.

In their experiments, the scientists were able to successfully insert their new plasmid into human cells in the lab.

These alterations to the standard CRISPR process could mean scientists will be able to do more widespread gene editing. The genes and proteins inside cells interact in incredibly complex ways, and sometimes just making one cut or change at a time can be limiting.

For example, the new technique could mean the activity of certain genes can be increased at the same time as the activity of other genes gets reduced – and all of these operations can be scheduled with greater precision too.

There's a catch here, however. We don't know exactly how more changes might play out in the organism being edited. As we've seen in the past, there may be unexpected secondary changes, and the more genes you change, the higher the risk.

"Direct repeat sequences and spacers containing complementary sequences [..] could generate complex secondary RNA structures affecting the maturation of CRISPR RNAs in cells," the team writes in their paper

"Consequently, complementary regions in pre-CRISPR RNA must be considered to improve CRISPR RNA maturation. Future work overcoming these limitations will open up numerous applications for highly multiplexed genome engineering."

We've already seen CRISPR used to cut out genes responsible for disease, and to kill off superbugs. Scientists say there's much more to come, and they now have an even more versatile and comprehensive toolkit at their disposal.

"Our method provides a powerful platform to investigate and orchestrate the sophisticated genetic programs underlying complex cell behaviours," the team writes.

The research has been published in Nature Methods.