Expanding the target range of CRISPR/Cas systems

University of Toronto researchers have created a genome-editing technique that allows small changes in target DNA but retains local specificity, which could help realize the potential of CRISPR/Cas-based gene therapy and pathogen diagnosis

Scientists program current CRISPR/Cas systems to recognize and cut precise DNA sequences to avoid effects such as cutting the wrong sequence or encouraging unwanted mutations

Basil Hubbard, lead researcher on the study, said: "There has been a lot of work to make CRISPR/Cas systems more specific

The discovery was recently published in the journal Nature Communications

The CRISPR-Cas system consists of two main molecules: a CRISPR guide-RNA, which contains nucleotide base pairs (various combinations of adenine, cytosine, guanine, and thymine) that guide the system to find complementary DNA segments; and a Cas enzyme, which cuts DNA in order to manipulate other genetic codes

The new method replaces one or more of the four bases that make up the CRISPR guide-RNA with a universal base

"It acts like an asterisk or wildcard in a numeric search, in areas where we expect variation or no data," Hubbard said

Experimental CRISPR therapies have shown the potential to eliminate genetic diseases, including sickle cell anemia and muscular dystrophy

Hubbard said his team's approach could help with this, but so far it works best in vitro, requiring faster operation in a cellular context and perhaps a redesigned Cas enzyme

The technology has broad application prospects in the field of diagnosis

"There is tremendous diversity among pathogens, especially viruses and bacteria, which evolve very quickly," Hubbard said

Hubbard came up with the idea of ​​using universal bases while working to make the CRISPR/Cas system more specific

Off-target effects of removing the wrong gene can have adverse effects on a patient's health, including the development of cancer

"Our current study gives us more options for tailoring CRISPR specificity," Hubbard said

Hubbard's lab has filed a patent for the technology and is seeking to partner with a company that specializes in CRISPR diagnostics
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He hopes the system will help provide a fast, accurate and affordable way to diagnose several diseases, including COVID-19
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Guide RNAs containing universal bases enable Cas9/Cas12a recognition of polymorphic sequences