Efficient Targeted Knock-In in Non-Dividing Cells Using Engineered Nucleases
Homology-independent targeted integration strategy that allows for an efficient targeted knock-in in both in vitro and in vivo
Background
Recent advances in targeted genome editing have transformed biomedical research. With the advent of the CRISPR/Cas9 system, genome editing has become faster, cheaper, and more accurate, and is now an invaluable tool for researchers in the biological sciences. Although this technology has been revolutionary for the research community and holds great potential for gene therapy, it is not an efficient tool for the targeted integration of transgenes in non-dividing cells. Currently, site-specific transgene integration using CRISPR/Cas9 exploits the homology-directed repair (HDR) pathway, which is inefficient in primary cell types and only functions in actively dividing cells. This limits the utility of CRISPR/Cas9 and other engineered nucleases in non-dividing cells, which are the major constituents of adult tissues.
Technology Overview
Investigators at Salk have developed a homology-independent targeted integration strategy that overcomes these limitations. This new strategy allows for an efficient targeted knock-in in both dividing and non-dividing cells in vitro and more importantly, in vivo. This innovative approach will not only help to advance basic biological research but has the potential to make targeted gene therapy safer and more efficient.
Further Details:
- Suzuki et. al. In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration Nature. Published online 16 November 2016. doi:10.1038/nature20565
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Stage of Development
Proof of concept in a rat model of retinitis pigmentosa.
Benefits
- Works in both dividing and non-dividing cells
- Applicable to in vitro and in vivo systems
- Only targeted gene insertion methods applicable to non-dividing cells
- Increased efficiencies compared to traditional methods
Applications
- In vivo targeted gene-replacement therapy
- Tissue and animal disease models
- Cellular engineering (stem cells, CAR-T cells)
- Live tracking of post-mitotic cells