4.8 Article

CRISPR prime editing with ribonucleoprotein complexes in zebrafish and primary human cells

Journal

NATURE BIOTECHNOLOGY
Volume 40, Issue 2, Pages 189-+

Publisher

NATURE PORTFOLIO
DOI: 10.1038/s41587-021-00901-y

Keywords

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Funding

  1. Hassenfeld Scholar Award
  2. NIH [R35 GM118158, R01 GM134069, RM1 HG009490]
  3. National Human Genome Research Institute Genomic Innovator Award [R35 HG010717]
  4. Defense Advanced Research Projects Agency Safe Genes program [HR0011-17-2-0042]
  5. Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) [417577129]
  6. China Scholarship Council [201808210354]
  7. John Hansen Research Grant from DKMS [DKMS-SLS-JHRG-2020-04]

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This study demonstrated prime editing using purified ribonucleoprotein complexes, achieving desired edits in zebrafish embryos and human cells. However, unintended insertions, deletions, and guide RNA scaffold incorporations were also observed.
Prime editors have been delivered using DNA or RNA vectors. Here we demonstrate prime editing with purified ribonucleoprotein complexes. We introduced somatic mutations in zebrafish embryos with frequencies as high as 30% and demonstrate germline transmission. We also observed unintended insertions, deletions and prime editing guide RNA (pegRNA) scaffold incorporations. In HEK293T and primary human T cells, prime editing with purified ribonucleoprotein complexes introduced desired edits with frequencies of up to 21 and 7.5%, respectively.

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