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| Item Type: | Article |
|---|---|
| Title: | Efficient generation of Rosa26 knock-in mice using CRISPR/Cas9 in C57BL/6 zygotes |
| Creators: |
Chu, V.T.  ORCID: https://orcid.org/0000-0003-1327-8696, Weber, T. ORCID: https://orcid.org/0000-0003-2079-842X, Graf, R. ORCID: https://orcid.org/0000-0002-5400-9938, Sommermann, T. ORCID: https://orcid.org/0000-0003-2899-0256, Petsch, K., Sack, U., Volchkov, P., Rajewsky, K. ORCID: https://orcid.org/0000-0002-6633-6370 and Kühn, R. ORCID: https://orcid.org/0000-0003-1694-9803
|
| Abstract: | BACKGROUND: The CRISPR/Cas9 system is increasingly used for gene inactivation in mouse zygotes, but homology-directed mutagenesis and use of inbred embryos are less established. In particular, Rosa26 knock-in alleles for the insertion of transgenes in a genomic 'safe harbor' site, have not been produced. Here we applied CRISPR/Cas9 for the knock-in of 8-11 kb inserts into Rosa26 of C57BL/6 zygotes. RESULTS: We found that 10-20 % of live pups derived from microinjected zygotes were founder mutants, without apparent off-target effects, and up to 50 % knock-in embryos were recovered upon coinjection of Cas9 mRNA and protein. Using this approach, we established a new mouse line for the Cre/loxP-dependent expression of Cas9. CONCLUSIONS: Altogether, our protocols and resources support the fast and direct generation of new Rosa26 knock-in alleles and of Cas9-mediated in vivo gene editing in the widely used C57BL/6 inbred strain. |
| Keywords: | CRISPR, Cas9, Knock-In Mice, Rosa26, Zygotes, Animals, Mice |
| Source: | BMC Biotechnology |
| ISSN: | 1472-6750 |
| Publisher: | BioMed Central |
| Volume: | 16 |
| Number: | 1 |
| Page Range: | 4 |
| Date: | 16 January 2016 |
| Official Publication: | https://doi.org/10.1186/s12896-016-0234-4 |
| PubMed: | View item in PubMed |
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