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| Item Type: | Article |
|---|---|
| Title: | Increasing the efficiency of homology-directed repair for CRISPR-Cas9-induced precise gene editing in mammalian cells |
| Creators Name: | Chu, V.T., Weber, T., Wefers, B., Wurst, W., Sander, S., Rajewsky, K. and Kühn, R. |
| Abstract: | The insertion of precise genetic modifications by genome editing tools such as CRISPR-Cas9 is limited by the relatively low efficiency of homology-directed repair (HDR) compared with the higher efficiency of the nonhomologous end-joining (NHEJ) pathway. To enhance HDR, enabling the insertion of precise genetic modifications, we suppressed the NHEJ key molecules KU70, KU80 or DNA ligase IV by gene silencing, the ligase IV inhibitor SCR7 or the coexpression of adenovirus 4 E1B55K and E4orf6 proteins in a 'traffic light' and other reporter systems. Suppression of KU70 and DNA ligase IV promotes the efficiency of HDR 4-5-fold. When co-expressed with the Cas9 system, E1B55K and E4orf6 improved the efficiency of HDR up to eightfold and essentially abolished NHEJ activity in both human and mouse cell lines. Our findings provide useful tools to improve the frequency of precise gene modifications in mammalian cells. |
| Keywords: | Adenoviridae, Adenovirus E4 Proteins, CRISPR-Cas Systems, Cell Line, DNA End-Joining Repair, DNA Ligases, Double-Stranded DNA Breaks, Gene Expression Regulation, Genetic Engineering, Homologous Recombination, Human Genome, Viral Proteins, Animals, Mice |
| Source: | Nature Biotechnology |
| ISSN: | 1087-0156 |
| Publisher: | Nature Publishing Group |
| Volume: | 33 |
| Number: | 5 |
| Page Range: | 543-548 |
| Date: | May 2015 |
| Additional Information: | Erratum in: Nat Biotechnol 36(2): 196. |
| Official Publication: | https://doi.org/10.1038/nbt.3198 |
| PubMed: | View item in PubMed |
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