Targeting fidelity of adenine and cytosine base editors in mouse embryos

Hye Kyung Lee; Michaela Willi; Shannon M Miller; Sojung Kim; Chengyu Liu; David R Liu; Lothar Hennighausen

doi:10.1038/s41467-018-07322-7

Targeting fidelity of adenine and cytosine base editors in mouse embryos

Nat Commun. 2018 Nov 15;9(1):4804. doi: 10.1038/s41467-018-07322-7.

Authors

Hye Kyung Lee¹, Michaela Willi², Shannon M Miller^{3

4

5}, Sojung Kim², Chengyu Liu⁶, David R Liu^{3

4

5}, Lothar Hennighausen⁷

Affiliations

¹ Laboratory of Genetics and Physiology, National Institute of Diabetes and Digestive and Kidney Diseases, US National Institutes of Health, Bethesda, MD, 20892, USA. hyekyung.lee@nih.gov.
² Laboratory of Genetics and Physiology, National Institute of Diabetes and Digestive and Kidney Diseases, US National Institutes of Health, Bethesda, MD, 20892, USA.
³ Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of MIT and Harvard, Cambridge, MA, 02141, USA.
⁴ Howard Hughes Medical Institute, Harvard University, Cambridge, MA, 02138, USA.
⁵ Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, 02138, USA.
⁶ Transgenic Core, National Heart, Lung, and Blood Institute, US National Institutes of Health, Bethesda, MD, 20892, USA.
⁷ Laboratory of Genetics and Physiology, National Institute of Diabetes and Digestive and Kidney Diseases, US National Institutes of Health, Bethesda, MD, 20892, USA. lotharh@niddk.nih.gov.

Abstract

Base editing directly converts a target base pair into a different base pair in the genome of living cells without introducing double-stranded DNA breaks. While cytosine base editors (CBE) and adenine base editors (ABE) are used to install and correct point mutations in a wide range of organisms, the extent and distribution of off-target edits in mammalian embryos have not been studied in detail. We analyze on-target and proximal off-target editing at 13 loci by a variety of CBEs and ABE in more than 430 alleles generated from mouse zygotic injections using newly generated and published sequencing data. ABE predominantly generates anticipated A•T-to-G•C edits. Among CBEs, SaBE3 and BE4, result in the highest frequencies of anticipated C•G-to-T•A products relative to editing byproducts. Together, these findings highlight the remarkable fidelity of ABE in mouse embryos and identify preferred CBE variants when fidelity in vivo is critical.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Adenine / metabolism*
Alleles
Animals
Animals, Genetically Modified
CRISPR-Associated Protein 9 / genetics
CRISPR-Associated Protein 9 / metabolism
CRISPR-Cas Systems*
Cytosine / metabolism*
Embryo, Mammalian
Gene Editing / methods*
Genetic Loci
Mice
Microinjections
Point Mutation*
RNA, Guide, CRISPR-Cas Systems
Reproducibility of Results
Sensitivity and Specificity
Zygote

Substances

Cytosine
CRISPR-Associated Protein 9
Adenine

Abstract

Publication types

MeSH terms

Substances

Grants and funding