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Prime editing primarily induces undesired outcomes in mice

Tomomi Aida, View ORCID ProfileJonathan J. Wilde, Lixin Yang, Yuanyuan Hou, Mengqi Li, Dongdong Xu, Jianbang Lin, Peimin Qi, Zhonghua Lu, Guoping Feng
doi: https://doi.org/10.1101/2020.08.06.239723
Tomomi Aida
1McGovern Institute for Brain Research, Department of Brain & Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA
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  • For correspondence: aidat@mit.edu wildej@mit.edu zh.lu@siat.ac.cn fengg@mit.edu
Jonathan J. Wilde
1McGovern Institute for Brain Research, Department of Brain & Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA
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  • For correspondence: aidat@mit.edu wildej@mit.edu zh.lu@siat.ac.cn fengg@mit.edu
Lixin Yang
3Shenzhen Key Laboratory for Molecular Biology of Neural Development, Guangdong Provincial Key Laboratory of Brain Connectome and Behavior, CAS Key Laboratory of Brain Connectome and Manipulation, The Brain Cognition and Brain Disease Institute, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; Shenzhen-Hong Kong Institute of Brain Science, Shenzhen, China
4University of Chinese Academy of Sciences, Beijing, China
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Yuanyuan Hou
1McGovern Institute for Brain Research, Department of Brain & Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA
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Mengqi Li
3Shenzhen Key Laboratory for Molecular Biology of Neural Development, Guangdong Provincial Key Laboratory of Brain Connectome and Behavior, CAS Key Laboratory of Brain Connectome and Manipulation, The Brain Cognition and Brain Disease Institute, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; Shenzhen-Hong Kong Institute of Brain Science, Shenzhen, China
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Dongdong Xu
3Shenzhen Key Laboratory for Molecular Biology of Neural Development, Guangdong Provincial Key Laboratory of Brain Connectome and Behavior, CAS Key Laboratory of Brain Connectome and Manipulation, The Brain Cognition and Brain Disease Institute, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; Shenzhen-Hong Kong Institute of Brain Science, Shenzhen, China
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Jianbang Lin
3Shenzhen Key Laboratory for Molecular Biology of Neural Development, Guangdong Provincial Key Laboratory of Brain Connectome and Behavior, CAS Key Laboratory of Brain Connectome and Manipulation, The Brain Cognition and Brain Disease Institute, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; Shenzhen-Hong Kong Institute of Brain Science, Shenzhen, China
4University of Chinese Academy of Sciences, Beijing, China
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Peimin Qi
1McGovern Institute for Brain Research, Department of Brain & Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA
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Zhonghua Lu
3Shenzhen Key Laboratory for Molecular Biology of Neural Development, Guangdong Provincial Key Laboratory of Brain Connectome and Behavior, CAS Key Laboratory of Brain Connectome and Manipulation, The Brain Cognition and Brain Disease Institute, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; Shenzhen-Hong Kong Institute of Brain Science, Shenzhen, China
4University of Chinese Academy of Sciences, Beijing, China
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  • For correspondence: aidat@mit.edu wildej@mit.edu zh.lu@siat.ac.cn fengg@mit.edu
Guoping Feng
1McGovern Institute for Brain Research, Department of Brain & Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA
2Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA
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  • For correspondence: aidat@mit.edu wildej@mit.edu zh.lu@siat.ac.cn fengg@mit.edu
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Summary

Genome editing has transformed biomedical science, but is still unpredictable and often induces undesired outcomes. Prime editing (PE) is a promising new approach due to its proposed flexibility and ability to avoid unwanted indels. Here, we show highly efficient PE-mediated genome editing in mammalian zygotes. Utilizing chemically modified guideRNAs, PE efficiently introduced 10 targeted modifications including substitutions, deletions, and insertions across 6 genes in mouse embryos. However, we unexpectedly observed a high frequency of undesired outcomes such as large deletions and found that these occurred more often than pure intended edits across all of the edits/genes. We show that undesired outcomes result from the double-nicking PE3 strategy, but that omission of the second nick largely ablates PE function. However, sequential double-nicking with PE3b, which is only applicable to a fraction of edits, eliminated undesired outcomes. Overall, our findings demonstrate the promising potential of PE for predictable, flexible, and highly efficient in vivo genome editing, but highlight the need for improved variations of PE before it is ready for widespread use.

Competing Interest Statement

The authors have declared no competing interest.

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Posted August 06, 2020.
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Prime editing primarily induces undesired outcomes in mice
Tomomi Aida, Jonathan J. Wilde, Lixin Yang, Yuanyuan Hou, Mengqi Li, Dongdong Xu, Jianbang Lin, Peimin Qi, Zhonghua Lu, Guoping Feng
bioRxiv 2020.08.06.239723; doi: https://doi.org/10.1101/2020.08.06.239723
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Prime editing primarily induces undesired outcomes in mice
Tomomi Aida, Jonathan J. Wilde, Lixin Yang, Yuanyuan Hou, Mengqi Li, Dongdong Xu, Jianbang Lin, Peimin Qi, Zhonghua Lu, Guoping Feng
bioRxiv 2020.08.06.239723; doi: https://doi.org/10.1101/2020.08.06.239723

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