CRISPR-Cas9 Knock-In Efficiency Enhanced Using Small-Molecule Inhibitor Pair, Study Shows
Wisconsin Eagle/10242338

MADISON, Wis. ~ A new study published in Nature Communications has demonstrated how small molecule inhibitors can be used to improve the precision and efficiency of CRISPR-Cas9 gene editing. The research, led by scientists from AstraZeneca, was conducted with the help of Promega research scientists Marie Schwinn and Michael Slater.

CRISPR-Cas9 is a genome editing technique that has revolutionized biomedical research since it was first unveiled in 2012. It enables researchers to cut DNA at specific sites and insert or remove genetic material, allowing for a level of genetic manipulation that was previously impossible.

The authors analyzed a library of 20,548 small molecules to develop strategies for improving this gene editing technique. They identified two inhibitors targeting the proteins DNA-dependent Protein Kinase (DNA-PK) and DNA Polymerase Theta (PolΘ) and demonstrated that the combination of these two molecules dramatically increased the performance of "knock-in" gene editing. This strategy, dubbed 2iHDR, outperformed previously described strategies that aimed to increase knock-in efficiency with small molecule inhibitors.

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The paper also documents a new system based on next-generation sequencing (NGS) that can be used to characterize the effects of inhibitors on CRISPR-Cas9. This open-access tool, which the authors named KI-Seq, enables researchers to analyze the contributions of different DNA repair pathways following CRISPR-Cas9 treatment. Unlike comparable tools, KI-Seq can be used with any cell type, including non-dividing primary cells.

The findings from this study are key to advancing the use of CRISPR techniques in research and clinical applications using a wide range of cell lines. The paper titled "Simultaneous inhibition of DNA-PK and Polϴ improves integration efficiency and precision of genome editing" provides further information on this breakthrough discovery. Promega technologies that can be used with CRISPR-Cas9 are also available at www.promega.com/CRISPR for those interested in learning more about this technology.

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