PUBLICATION
Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction
- Authors
- Cornean, A., Gierten, J., Welz, B., Mateo, J.L., Thumberger, T., Wittbrodt, J.
- ID
- ZDB-PUB-220405-21
- Date
- 2022
- Source
- eLIFE 11: (Journal)
- Registered Authors
- Wittbrodt, Jochen
- Keywords
- developmental biology, genetics, genomics, zebrafish
- MeSH Terms
-
- Animals
- Adenine
- DNA
- Zebrafish*/genetics
- Gene Editing*
- Cytosine
- Mutation
- CRISPR-Cas Systems
- PubMed
- 35373735 Full text @ Elife
Citation
Cornean, A., Gierten, J., Welz, B., Mateo, J.L., Thumberger, T., Wittbrodt, J. (2022) Precise in vivo functional analysis of DNA variants with base editing using ACEofBASEs target prediction. eLIFE. 11:.
Abstract
Single nucleotide variants (SNVs) are prevalent genetic factors shaping individual trait profiles and disease susceptibility. The recent development and optimizations of base editors, rubber and pencil genome editing tools now promise to enable direct functional assessment of SNVs in model organisms. However, the lack of bioinformatic tools aiding target prediction limits the application of base editing in vivo. Here, we provide a framework for adenine and cytosine base editing in medaka (Oryzias latipes) and zebrafish (Danio rerio), ideal for scalable validation studies. We developed an online base editing tool ACEofBASEs (a careful evaluation of base-edits), to facilitate decision-making by streamlining sgRNA design and performing off-target evaluation. We used state-of-the-art adenine (ABE) and cytosine base editors (CBE) in medaka and zebrafish to edit eye pigmentation genes and transgenic GFP function with high efficiencies. Base editing in the genes encoding troponin T and the potassium channel ERG faithfully recreated known cardiac phenotypes. Deep-sequencing of alleles revealed the abundance of intended edits in comparison to low levels of insertion or deletion (indel) events for ABE8e and evoBE4max. We finally validated missense mutations in novel candidate genes of congenital heart disease (CHD) dapk3, ube2b, usp44, and ptpn11 in F0 and F1 for a subset of these target genes with genotype-phenotype correlation. This base editing framework applies to a wide range of SNV-susceptible traits accessible in fish, facilitating straight-forward candidate validation and prioritization for detailed mechanistic downstream studies.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping