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Publication . Article . 2020

Efficient Generation and Correction of Mutations in Human iPS Cells Utilizing mRNAs of CRISPR Base Editors and Prime Editors

Duran Sürün; Aksana D. Schneider; Jovan Mircetic; Katrin Neumann; Felix Lansing; Maciej Paszkowski-Rogacz; Vanessa Hänchen; +2 Authors
Open Access  
Published: 06 May 2020
Publisher: Zenodo
Abstract In contrast to CRISPR/Cas9 nucleases, CRISPR base editors (BE) and prime editors (PE) enable predefined nucleotide exchanges in genomic sequences without generating DNA double strand breaks. Here, we employed BE and PE mRNAs in conjunction with chemically synthesized sgRNAs and pegRNAs for efficient editing of human induced pluripotent stem cells (iPSC). Whereas we were unable to correct a disease-causing mutation in patient derived iPSCs using a CRISPR/Cas9 nuclease approach, we corrected the mutation back to wild type with high efficiency utilizing an adenine BE. We also used adenine and cytosine BEs to introduce nine different cancer associated TP53 mutations into human iPSCs with up to 90% efficiency, generating a panel of cell lines to investigate the biology of these mutations in an isogenic background. Finally, we pioneered the use of prime editing in human iPSCs, opening this important cell type for the precise modification of nucleotides not addressable by BEs and to multiple nucleotide exchanges. These approaches eliminate the necessity of deriving disease specific iPSCs from human donors and allows the comparison of different disease-causing mutations in isogenic genetic backgrounds.
Subjects by Vocabulary

Library of Congress Subject Headings: lcsh:Genetics lcsh:QH426-470

Microsoft Academic Graph classification: Induced pluripotent stem cell Cas9 Biology Mutation medicine.disease_cause medicine Computational biology DNA chemistry.chemical_compound chemistry Wild type Nuclease biology.protein CRISPR Cytosine


CRISPR/Cas9, base editors, prime editors, human induced pluripotent stem cells, mRNA, Genetics (clinical), Genetics, Article

Funded by
Designer recombinases for efficient and safe genome surgery
  • Funder: European Commission (EC)
  • Project Code: 742133
  • Funding stream: H2020 | ERC | ERC-ADG
Validated by funder
Unlocking Precision Gene Therapy
  • Funder: European Commission (EC)
  • Project Code: 825825
  • Funding stream: H2020 | RIA
Validated by funder
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