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CRISPR-STOP: gene silencing through base-editing-induced nonsense mutations

Nature Methods volume 14, pages 710712 (2017) | Download Citation

Abstract

CRISPR–Cas9-induced DNA damage may have deleterious effects at high-copy-number genomic regions. Here, we use CRISPR base editors to knock out genes by changing single nucleotides to create stop codons. We show that the CRISPR-STOP method is an efficient and less deleterious alternative to wild-type Cas9 for gene-knockout studies. Early stop codons can be introduced in 17,000 human genes. CRISPR-STOP-mediated targeted screening demonstrates comparable efficiency to WT Cas9, which indicates the suitability of our approach for genome-wide functional screenings.

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Acknowledgements

This research was funded through a pilot project (NIDDK P50 DK096373), V Scholar award to M.A. from V Cancer Research Foundation, and UVA Cancer Center pilot project awards to M.A. We would like to thank S. Shang for his constructive feedback and critical reading of the manuscript.

Author information

Affiliations

  1. Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia, USA.

    • Cem Kuscu
    • , Mahmut Parlak
    • , Turan Tufan
    • , Jiekun Yang
    • , Karol Szlachta
    • , Xiaolong Wei
    • , Rashad Mammadov
    •  & Mazhar Adli

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Contributions

C.K. and M.A. designed the study; C.K. performed experiments. J.Y. and K.S. performed computational work; and T.T., M.P., X.W. and R.M. helped C.K. with experiments.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Mazhar Adli.

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DOI

https://doi.org/10.1038/nmeth.4327

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