PUBLICATION
Optimizing Prime Editing in Zebrafish
- Authors
- Basharat, R., Rizzo, G., Zoodsma, J.D., Wollmuth, L.P., Sirotkin, H.I.
- ID
- ZDB-PUB-250920-1
- Date
- 2025
- Source
- The CRISPR journal 8: 426-435 (Journal)
- Registered Authors
- Sirotkin, Howard
- Keywords
- none
- MeSH Terms
-
- Animals
- CRISPR-Cas Systems*/genetics
- Clustered Regularly Interspaced Short Palindromic Repeats/genetics
- Gene Editing*/methods
- Mutation, Missense
- RNA, Guide, CRISPR-Cas Systems/genetics
- RNA, Messenger/genetics
- Zebrafish*/genetics
- PubMed
- 40971219 Full text @ CRISPR J
Citation
Basharat, R., Rizzo, G., Zoodsma, J.D., Wollmuth, L.P., Sirotkin, H.I. (2025) Optimizing Prime Editing in Zebrafish. The CRISPR journal. 8:426-435.
Abstract
Prime editing is a clustered regularly interspaced short palindromic repeats-based approach that enables the introduction of precise genetic modifications, including missense mutations, making it valuable for generating disease models. The comparative performance of novel prime editor (PE) variants in zebrafish remains largely unexplored. Here, we systematically evaluated the efficiency of five PEs-PE2, PE6b, PE6c, PEmax, and PE7-in zebrafish. We tested mRNA encoding for each of these PEs with prime editing guide RNAs (pegRNAs) designed to install five missense mutations. Efficient editing was achieved at four of the five sites with multiple PEs. Among these, PEmax emerged as the most efficient editor for introducing pure prime edits, with rates reaching 15.34%. We found that strategies proposed to block 3' degradation of pegRNAs (epegRNAs and addition of a La RNA binding motif to the PE) did not improve performance in our assays. Together, these findings establish PEmax as a robust tool to introduce missense mutations into zebrafish.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping