New method for improving success of gene editing technique

A team of researchers led by a UW-Madison biomedical engineering professor has published a method for significantly improving the success of gene editing with CRISPR-Cas9 technology.

CRISPR has been widely adopted by geneticists because it’s much cheaper and easier than previous methods for gene editing. It works in a cut-and-paste fashion, allowing scientists to target specific parts of the genome to be altered, resulting in a change in biological expression of that portion of the genetic code.

While the cutting part of the process is very consistent, making desired changes has been less so, with error rates as high as 50 percent, according to a release from UW-Madison.

The new method improves the chance of correctly rewriting the DNA sequence by a factor of 10, thanks to the addition of a glue-like molecule called RNA aptamer. Using this RNA aptamer, the team led by Krishanu Saha has created a method for delivering a complete CRISPR ‘repair kit’ to the target site.

“The kit provides not only the molecular scissors, but also the correct template for the cell machinery to fix the DNA cut with,” Saha said. “Since the RNA aptamer is strong and very stable, everything we need is getting to the right place within the cell in one fell swoop.”

The UW-Madison release highlights two main advantages of Saha’s method: the kit has only non-viral reagents, which simplifies the manufacturing process and reduces concerns about safety; and attaching the RNA aptamer is ‘much easier’ than modifying the actual Cas9 protein and provides more flexibility.

“We can add other biomolecules to this kit, much like you would click an extra Lego block into an already existing structure,” said Jared Carlson-Stevermer, another author on the paper and a grad student in Saha’s lab. Adding a fluorescent tag to modified cells for quick identification, for example, could lead to an accuracy rate of over 98 percent, Saha says.

“There is no shortage of candidates for this kind of genetic surgery, as tens of thousands of diseases are due to small sequence errors that could be fixed with this technology,” Saha said. “Our next goal is to test the method in animal models and work on writing longer stretches of DNA.”

The study was funded by the National Institutes of Health and the National Science Foundation. Some of the co-authors have filed a patent application with the Wisconsin Alumni Research Foundation.