Evolving high fidelity CRISPR

ANNA CERESETO BISTURI GENOMICO BIOMEDICINA

credit Alessio Coser

It’s called evoCas9, and it’s the most accurate CRISPR editing system yet, according to a study just published in Nature Biotechnology. Researchers at the University of Trento, in northern Italy, induced random mutations in vitro on a piece of a bacterial gene coding for the DNA-cutting enzyme (the REC3 domain of SpCas9) and then screened the mutated variants in vivo in yeast colonies by looking at their color. If the molecular scissors work properly, cutting only the right target, the yeast becomes red, but colonies are white if CRISPR cuts off target.

“We started working on it in 2015 when CRISPR was very young”, says Anna Cereseto. “In the meantime, this sector has exploded. It was a race against time “, adds Antonio Casini. Nobody in the team expected such an achievement: by combining four beneficial mutations, they got a variant whose fidelity exceeds both wild-type and rationally designed Cas9 variants, while maintaining near wild-type editing efficiency. “We report a reduction of almost 99% in off-target mutations. As far as we know, our Cas9 is the most specific ever developed”, researchers say. The same strategy can now be used to evolve further enzymes of the Cas family, including base-editors designed to replace DNA letters without even cutting. The University of Trento has already filed a patent application on evoCas9, and a start-up is due to launch soon.

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