Increasing the efficiency of precise genome editing with CRISPR-Cas9 by inhibition of nonhomologous end joining

Takeshi Maruyama, Stephanie K. Dougan, Matthias C. Truttmann, Angelina M. Bilate, Jessica R. Ingram, Hidde L. Ploegh

Research output: Contribution to journalArticle

459 Citations (Scopus)

Abstract

Methods to introduce targeted double-strand breaks (DSBs) into DNA enable precise genome editing by increasing the rate at which externally supplied DNA fragments are incorporated into the genome through homologous recombination. The efficiency of these methods is limited by nonhomologous end joining (NHEJ), an alternative DNA repair pathway that competes with homology-directed repair (HDR). To promote HDR at the expense of NHEJ, we targeted DNA ligase IV, a key enzyme in the NHEJ pathway, using the inhibitor Scr7. Scr7 treatment increased the efficiency of HDR-mediated genome editing, using Cas9 in mammalian cell lines and in mice for all four genes examined, up to 19-fold. This approach should be applicable to other customizable endonucleases, such as zinc finger nucleases and transcription activator-like effector nucleases, and to nonmammalian cells with sufficiently conserved mechanisms of NHEJ and HDR.

Original languageEnglish
Pages (from-to)538-542
Number of pages5
JournalNature Biotechnology
Volume33
Issue number5
DOIs
Publication statusPublished - 2015 May 12
Externally publishedYes

Fingerprint

DNA End-Joining Repair
Clustered Regularly Interspaced Short Palindromic Repeats
Joining
Repair
Genes
DNA
Double-Stranded DNA Breaks
Endonucleases
Homologous Recombination
Zinc Fingers
Genome
Cell Line
DNA Ligases
Enzymes
Transcription
Zinc
Cells
Gene Editing

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Biomedical Engineering
  • Molecular Medicine

Cite this

Increasing the efficiency of precise genome editing with CRISPR-Cas9 by inhibition of nonhomologous end joining. / Maruyama, Takeshi; Dougan, Stephanie K.; Truttmann, Matthias C.; Bilate, Angelina M.; Ingram, Jessica R.; Ploegh, Hidde L.

In: Nature Biotechnology, Vol. 33, No. 5, 12.05.2015, p. 538-542.

Research output: Contribution to journalArticle

Maruyama, Takeshi ; Dougan, Stephanie K. ; Truttmann, Matthias C. ; Bilate, Angelina M. ; Ingram, Jessica R. ; Ploegh, Hidde L. / Increasing the efficiency of precise genome editing with CRISPR-Cas9 by inhibition of nonhomologous end joining. In: Nature Biotechnology. 2015 ; Vol. 33, No. 5. pp. 538-542.
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