Improved gRNA secondary structures allow editing of target sites resistant to CRISPR-Cas9 cleavage
Stephan Riesenberg (),
Nelly Helmbrecht,
Philipp Kanis,
Tomislav Maricic and
Svante Pääbo
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Stephan Riesenberg: Max Planck Institute for Evolutionary Anthropology
Nelly Helmbrecht: Max Planck Institute for Evolutionary Anthropology
Philipp Kanis: Max Planck Institute for Evolutionary Anthropology
Tomislav Maricic: Max Planck Institute for Evolutionary Anthropology
Svante Pääbo: Max Planck Institute for Evolutionary Anthropology
Nature Communications, 2022, vol. 13, issue 1, 1-8
Abstract:
Abstract The first step in CRISPR-Cas9-mediated genome editing is the cleavage of target DNA sequences that are complementary to so-called spacer sequences in CRISPR guide RNAs (gRNAs). However, some DNA sequences are refractory to CRISPR-Cas9 cleavage, which is at least in part due to gRNA misfolding. To overcome this problem, we have engineered gRNAs with highly stable hairpins in their constant parts and further enhanced their stability by chemical modifications. The ‘Genome-editing Optimized Locked Design’ (GOLD)-gRNA increases genome editing efficiency up to around 1000-fold (from 0.08 to 80.5%) with a mean increase across different other targets of 7.4-fold. We anticipate that this improved gRNA will allow efficient editing regardless of spacer sequence composition and will be especially useful if a desired genomic site is difficult to edit.
Date: 2022
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-28137-7
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DOI: 10.1038/s41467-022-28137-7
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