Journal
SCIENTIFIC REPORTS
Volume 12, Issue 1, Pages -Publisher
NATURE PORTFOLIO
DOI: 10.1038/s41598-022-15972-3
Keywords
-
Categories
Funding
- Takeda Foundation
- Mitsubishi Foundation
- Nagase Science and Technology Foundation
- Ono Medical foundation
- Daiichi Sankyo Foundation
- AMED [21gm5010001h0005]
- Joint Research of the Exploratory Research Center on Life and Living Systems (ExCELLS)
- NIBB Collaborative Research Program
- JST FOREST
- JSPS [20J15808]
- [21H05287]
- [19H03412]
- [22H02820]
- [20H05791]
- [21H05148]
- [19K06673]
- [22H04648]
- [21H00427]
- [20H05311]
- [20H04922]
- [20K15701]
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Researchers have developed a new CRISPR/Cas9 method for rapid generation of knockout and transgenic African turquoise killifish. They have also successfully developed a method for knock-in of reporter genes without crossing. These advances will accelerate aging and developmental studies using the fish model.
The African turquoise killifish Nothobranchius furzeri (N. furzeri) is a useful model organism for studying aging, age-related diseases, and embryonic diapause. CRISPR/Cas9-mediated gene knockout and Tol2 transposon-mediated transgenesis in N. furzeri have been reported previously. However, these methods take time to generate knockout and transgenic fish. In addition, knock-in technology that inserts large DNA fragments as fluorescent reporter constructs into the target gene in N. furzeri has not yet been established. Here, we show that triple-target CRISPR-mediated single gene disruption efficiently produces whole-body biallelic knockout and enables the examination of gene function in the F0 generation. In addition, we developed a method for creating the knock-in reporter N. furzeri without crossing by optimizing the CRISPR/Cas9 system. These methods drastically reduce the duration of experiments, and we think that these advances will accelerate aging and developmental studies using N. furzeri.
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