Boosting targeted genome editing using the hei-tag
- Heidelberg University, Germany
Abstract
Precise, targeted genome editing by CRISPR/Cas9 is key for basic research and translational approaches in model and non-model systems. While active in all species tested so far, editing efficiencies still leave room for improvement. The bacterial Cas9 needs to be efficiently shuttled into the nucleus as attempted by fusion with nuclear localization signals (NLSs). Additional peptide tags such as FLAG- or myc-tags are usually added for immediate detection or straight-forward purification. Immediate activity is usually granted by administration of pre-assembled protein/RNA complexes. We present the 'hei-tag (high efficiency-tag)' which boosts the activity of CRISPR/Cas genome editing tools already when supplied as mRNA. The addition of the hei-tag, a myc tag coupled to an optimized NLS via a flexible linker, to Cas9 or a C-to-T base editor dramatically enhances the respective targeting efficiency. This results in an increase in bi-allelic editing, yet reduction of allele variance, indicating an immediate activity even at early developmental stages. The hei-tag boost is active in model systems ranging from fish to mammals, including tissue culture applications. The simple addition of the hei-tag allows to instantly upgrade existing and potentially highly adapted systems as well as to establish novel highly efficient tools immediately applicable at the mRNA level.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files.
Article and author information
Author details
Tinatini Tavhelidse-Suck
Funding
Deutsche Forschungsgemeinschaft (CRC873,project A3)
- Joachim Wittbrodt
Deutsche Forschungsgemeinschaft (FOR2509 P10,WI 1824/9-1)
- Joachim Wittbrodt
Deutsche Forschungsgemeinschaft (CRC1118,project S03)
- Marc Freichel
ERC-SyG H2020 (NO 810172)
- Joachim Wittbrodt
Deutsche Forschungsgemeinschaft (3DMM2O, EXC 2082/1 Wittbrodt C3)
- Joachim Wittbrodt
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2022, Thumberger et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 3,659
- views
-
- 707
- downloads
-
- 19
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Citations by DOI
-
- 19
- citations for umbrella DOI https://doi.org/10.7554/eLife.70558
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Boosting targeted genome editing using the hei-tag
eLife 11:e70558.
https://doi.org/10.7554/eLife.70558
