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Harnessing noncanonical crRNA for highly efficient genome editing

Author

Listed:
  • Guanhua Xun

    (University of Illinois at Urbana-Champaign
    University of Illinois at Urbana-Champaign)

  • Zhixin Zhu

    (University of Illinois at Urbana-Champaign
    University of Illinois at Urbana-Champaign)

  • Nilmani Singh

    (University of Illinois at Urbana-Champaign)

  • Jingxia Lu

    (University of Illinois at Urbana-Champaign)

  • Piyush K. Jain

    (University of Florida)

  • Huimin Zhao

    (University of Illinois at Urbana-Champaign
    University of Illinois at Urbana-Champaign
    University of Illinois at Urbana-Champaign)

Abstract

The CRISPR-Cas12a system is more advantageous than the widely used CRISPR-Cas9 system in terms of specificity and multiplexibility. However, its on-target editing efficiency is typically much lower than that of the CRISPR-Cas9 system. Here we improved its on-target editing efficiency by simply incorporating 2-aminoadenine (base Z, which alters canonical Watson-Crick base pairing) into the crRNA to increase the binding affinity between crRNA and its complementary DNA target. The resulting CRISPR-Cas12a (named zCRISPR-Cas12a thereafter) shows an on-target editing efficiency comparable to that of the CRISPR-Cas9 system but with much lower off-target effects than the CRISPR-Cas9 system in mammalian cells. In addition, zCRISPR-Cas12a can be used for precise gene knock-in and highly efficient multiplex genome editing. Overall, the zCRISPR-Cas12a system is superior to the CRISPR-Cas9 system, and our simple crRNA engineering strategy may be extended to other CRISPR-Cas family members as well as their derivatives.

Suggested Citation

  • Guanhua Xun & Zhixin Zhu & Nilmani Singh & Jingxia Lu & Piyush K. Jain & Huimin Zhao, 2024. "Harnessing noncanonical crRNA for highly efficient genome editing," Nature Communications, Nature, vol. 15(1), pages 1-13, December.
    • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-48012-x
    DOI: 10.1038/s41467-024-48012-x
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