Homology-directed repair in rodent zygotes using Cas9 and TALEN engineered proteins

The generation of genetically-modified organisms has been revolutionized by the development of new genome editing technologies based on the use of gene-specific nucleases, such as meganucleases, ZFNs, TALENs and CRISPRs-Cas9 systems. The most rapid and cost-effective way to generate genetically-modi...

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Bibliographic Details
Published in:Scientific reports 2015-10, Vol.5 (1), p.14410-14410, Article 14410
Main Authors: Ménoret, Séverine, De Cian, Anne, Tesson, Laurent, Remy, Séverine, Usal, Claire, Boulé, Jean-Baptiste, Boix, Charlotte, Fontanière, Sandra, Crénéguy, Alison, Nguyen, Tuan H., Brusselle, Lucas, Thinard, Reynald, Gauguier, Dominique, Concordet, Jean-Paul, Cherifi, Yacine, Fraichard, Alexandre, Giovannangeli, Carine, Anegon, Ignacio
Format: Article
Language:English
Subjects:
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Animals
CRISPR-Cas Systems - genetics
Food and Nutrition
Genetically modified organisms
Genetics
Genome editing
Genomes
Homology
Human health and pathology
Humanities and Social Sciences
Life Sciences
Mice
Mice, Inbred C57BL
Microinjection
Microinjections
mRNA
multidisciplinary
Nuclease
Nucleic acids
Protein Engineering
Proteins
Rats
Rats, Sprague-Dawley
Recombinational DNA Repair - genetics
Science
Transcription activator-like effector nucleases
Zygote - physiology
Zygotes
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Summary:The generation of genetically-modified organisms has been revolutionized by the development of new genome editing technologies based on the use of gene-specific nucleases, such as meganucleases, ZFNs, TALENs and CRISPRs-Cas9 systems. The most rapid and cost-effective way to generate genetically-modified animals is by microinjection of the nucleic acids encoding gene-specific nucleases into zygotes. However, the efficiency of the procedure can still be improved. In this work we aim to increase the efficiency of CRISPRs-Cas9 and TALENs homology-directed repair by using TALENs and Cas9 proteins, instead of mRNA, microinjected into rat and mouse zygotes along with long or short donor DNAs. We observed that Cas9 protein was more efficient at homology-directed repair than mRNA, while TALEN protein was less efficient than mRNA at inducing homology-directed repair. Our results indicate that the use of Cas9 protein could represent a simple and practical methodological alternative to Cas9 mRNA in the generation of genetically-modified rats and mice as well as probably some other mammals.
ISSN:2045-2322
2045-2322
DOI:10.1038/srep14410