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Miniature type V-F CRISPR-Cas nucleases enable targeted DNA modification in cells

Author

Listed:
  • Greta Bigelyte

    (Vilnius University)

  • Joshua K. Young

    (Corteva Agriscience™)

  • Tautvydas Karvelis

    (Vilnius University)

  • Karolina Budre

    (Vilnius University)

  • Rimante Zedaveinyte

    (Vilnius University)

  • Vesna Djukanovic

    (Corteva Agriscience™)

  • Elizabeth Ginkel

    (Corteva Agriscience™)

  • Sushmitha Paulraj

    (Corteva Agriscience™)

  • Stephen Gasior

    (Corteva Agriscience™)

  • Spencer Jones

    (Corteva Agriscience™)

  • Lanie Feigenbutz

    (Corteva Agriscience™)

  • Grace St. Clair

    (Corteva Agriscience™)

  • Pierluigi Barone

    (Corteva Agriscience™)

  • Jennifer Bohn

    (Corteva Agriscience™)

  • Ananta Acharya

    (Corteva Agriscience™)

  • Gina Zastrow-Hayes

    (Corteva Agriscience™)

  • Selgar Henkel-Heinecke

    (Leipzig University)

  • Arunas Silanskas

    (Vilnius University)

  • Ralf Seidel

    (Leipzig University)

  • Virginijus Siksnys

    (Vilnius University)

Abstract

Class 2 CRISPR systems are exceptionally diverse, nevertheless, all share a single effector protein that contains a conserved RuvC-like nuclease domain. Interestingly, the size of these CRISPR-associated (Cas) nucleases ranges from >1000 amino acids (aa) for Cas9/Cas12a to as small as 400-600 aa for Cas12f. For in vivo genome editing applications, compact RNA-guided nucleases are desirable and would streamline cellular delivery approaches. Although miniature Cas12f effectors have been shown to cleave double-stranded DNA, targeted DNA modification in eukaryotic cells has yet to be demonstrated. Here, we biochemically characterize two miniature type V-F Cas nucleases, SpCas12f1 (497 aa) and AsCas12f1 (422 aa), and show that SpCas12f1 functions in both plant and human cells to produce targeted modifications with outcomes in plants being enhanced with short heat pulses. Our findings pave the way for the development of miniature Cas12f1-based genome editing tools.

Suggested Citation

  • Greta Bigelyte & Joshua K. Young & Tautvydas Karvelis & Karolina Budre & Rimante Zedaveinyte & Vesna Djukanovic & Elizabeth Ginkel & Sushmitha Paulraj & Stephen Gasior & Spencer Jones & Lanie Feigenbu, 2021. "Miniature type V-F CRISPR-Cas nucleases enable targeted DNA modification in cells," Nature Communications, Nature, vol. 12(1), pages 1-8, December.
    • Handle: RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-26469-4
    DOI: 10.1038/s41467-021-26469-4
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    References listed on IDEAS

    as
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    1. Shuqian Zhang & Liting Song & Bo Yuan & Cheng Zhang & Jixin Cao & Jinlong Chen & Jiayi Qiu & Yilin Tai & Jingqi Chen & Zilong Qiu & Xing-Ming Zhao & Tian-Lin Cheng, 2023. "TadA reprogramming to generate potent miniature base editors with high precision," Nature Communications, Nature, vol. 14(1), pages 1-12, December.

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