A panel of eGFP reporters for single base editing by APOBEC-Cas9 editosome complexes

A. St Martin, D. J. Salamango, A. A. Serebrenik, N. M. Shaban, W. L. Brown, R. S. Harris

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

The prospect of introducing a single C-to-T change at a specific genomic location has become feasible with APOBEC-Cas9 editing technologies. We present a panel of eGFP reporters for quantification and optimization of single base editing by APOBEC-Cas9 editosomes. Reporter utility is demonstrated by comparing activities of seven human APOBEC3 enzymes and rat APOBEC1 (BE3). APOBEC3A and RNA binding-defective variants of APOBEC3B and APOBEC3H display the highest single base editing efficiencies. APOBEC3B catalytic domain complexes also elicit the lowest frequencies of adjacent off-target events. However, unbiased deep-sequencing of edited reporters shows that all editosomes have some degree of local off-target editing. Thus, further optimization is required to generate true single base editors and the eGFP reporters described here have the potential to facilitate this process.

Original languageEnglish (US)
Article number497
JournalScientific reports
Volume9
Issue number1
DOIs
StatePublished - Dec 1 2019

Bibliographical note

Funding Information:
We thank Silvo Conticello for thoughtful discussions and David Liu for sharing BE3. These studies were supported by NIGMS R01 GM118000 and NIAID R37 AI064046. A.S. received partial salary support from NSF-GRFP 00039202 and D.J.S. from NIH T90DE022732. R.S.H. is the Margaret Harvey Schering Land Grant Chair for Cancer Research, a Distinguished McKnight University Professor, and an Investigator of the Howard Hughes Medical Institute.

Publisher Copyright:
© 2019, The Author(s).

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