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Generation of multi-gene knockout rabbits using the Cas9/gRNA system
Quanmei Yan1, Quanjun Zhang1, Huaqiang Yang1
1Key Laboratory of Regenerative Biology, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530 China.
Cell Regeneration (London, England)
|November 20, 2014
Summary
CRISPR gene editing is now efficient in rabbits, enabling single and multiple gene knockouts. This powerful tool allows for precise genetic modifications in rabbits for research purposes.
Area of Science:
- Genetics
- Molecular Biology
- Animal Biotechnology
Background:
- The clustered regularly interspaced short palindromic repeat (CRISPR)-associated system (Cas) is a proven gene-editing technology in model organisms.
- Application of CRISPR technology in rabbits has been limited, hindering genetic research in this species.
Purpose of the Study:
- To establish and evaluate the efficiency of CRISPR-Cas9 gene editing for single and multiple gene knockouts in rabbits.
- To demonstrate the utility of CRISPR technology for rapid genetic modification in rabbits.
Main Methods:
- Microinjection of Cas9 mRNA and guide RNA (gRNA) into pronuclear-stage rabbit embryos.
- Assessment of gene knockout efficiency for single, double, triple, and quintuple gene targets.
Main Results:
- Achieved 100% biallelic gene knockout efficiency for single (IL2rg) and double (IL2rg and RAG1) gene targets.
- Demonstrated high efficiency for simultaneous gene mutation: 100% for 3 genes (IL2rg, RAG1, RAG2) and 33.3% for 5 genes (IL2rg, RAG1, RAG2, TIKI1, ALB).
Conclusions:
- The Cas9/gRNA system is a highly efficient and rapid tool for gene editing in rabbits.
- This technology enables both single-gene and multi-gene editing in rabbits, facilitating advanced genetic research.
