Efficient CRISPR/Cas9 genome editing with low off-target effects in zebrafish

Development (Cambridge) - Tập 140 Số 24 - Trang 4982-4987 - 2013
Alexander Hruscha1, Peter Krawitz2,3,4, Alexandra Rechenberg1, Verena Heinrich3, Jochen Hecht2,4, Christian Haass5,1,6, Bettina Schmid1,6
1German Center for Neurodegenerative Diseases (DZNE), Schillerstrasse 44, 80336 Munich, Germany
2Berlin Brandenburg Center for Regenerative Therapies, Charité Universitätsmedizin, 13353 Berlin, Germany
3Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin, 13353 Berlin, Germany
4Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany
5Adolf-Butenandt-Institute, Biochemistry, Ludwig-Maximilians University Munich, Schillerstrasse 44, 80336 Munich, Germany.
6Munich Cluster for Systems Neurology (SyNergy), 80336 Munich, Germany

Tóm tắt

Gene modifications in animal models have been greatly facilitated through the application of targeted genome editing tools. The prokaryotic CRISPR/Cas9 type II genome editing system has recently been applied in cell lines and vertebrates. However, we still have very limited information about the efficiency of mutagenesis, germline transmission rates and off-target effects in genomes of model organisms. We now demonstrate that CRISPR/Cas9 mutagenesis in zebrafish is highly efficient, reaching up to 86.0%, and is heritable. The efficiency of the CRISPR/Cas9 system further facilitated the targeted knock-in of a protein tag provided by a donor oligonucleotide with knock-in efficiencies of 3.5-15.6%. Mutation rates at potential off-target sites are only 1.1-2.5%, demonstrating the specificity of the CRISPR/Cas9 system. The ease and efficiency of the CRISPR/Cas9 system with limited off-target effects make it a powerful genome engineering tool for in vivo studies.

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Development (Cambridge)

Tập 140 Số 24

4982-4987

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