Scientific reports, 2017-12, Vol.7 (1), p.17472-17472, Article 17472
2017
Details
- Autor(en) / Beteiligte
- Titel
- CRISPR/Cas9 microinjection in oocytes disables pancreas development in sheep
- Ist Teil von
- Scientific reports, 2017-12, Vol.7 (1), p.17472-17472, Article 17472
- Ort / Verlag
- England: Nature Publishing Group
- Erscheinungsjahr
- 2017
- Link zum Volltext
- Quelle
- Free E-Journal (出版社公開部分のみ）
- Beschreibungen/Notizen
- One of the ultimate goals of regenerative medicine is the generation of patient-specific organs from pluripotent stem cells (PSCs). Sheep are potential hosts for growing human organs through the technique of blastocyst complementation. We report here the creation of pancreatogenesis-disabled sheep by oocyte microinjection of CRISPR/Cas9 targeting PDX1, a critical gene for pancreas development. We compared the efficiency of target mutations after microinjecting the CRISPR/Cas9 system in metaphase II (MII) oocytes and zygote stage embryos. MII oocyte microinjection reduced lysis, improved blastocyst rate, increased the number of targeted bi-allelic mutations, and resulted in similar degree of mosaicism when compared to zygote microinjection. While the use of a single sgRNA was efficient at inducing mutated fetuses, the lack of complete gene inactivation resulted in animals with an intact pancreas. When using a dual sgRNA system, we achieved complete PDX1 disruption. This PDX1 fetus lacked a pancreas and provides the basis for the production of gene-edited sheep as a host for interspecies organ generation. In the future, combining gene editing with CRISPR/Cas9 and PSCs complementation could result in a powerful approach for human organ generation.
- Sprache
- Englisch
- Identifikatoren
- ISSN: 2045-2322eISSN: 2045-2322DOI: 10.1038/s41598-017-17805-0Titel-ID: cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_5727233
- Format
- –
- Schlagworte
- Animals, Animals, Genetically Modified, Complementation, Coumarins, CRISPR, CRISPR-Cas Systems, Embryos, Fetuses, Gene Editing - methods, Gene Knockdown Techniques - methods, Homeodomain Proteins - genetics, Homeodomain Proteins - metabolism, Lysis, Metaphase, Microinjection, Microinjections, Mosaicism, Mutation, Oocytes, Oocytes - metabolism, Ovis aries, Pancreas, Pancreas - embryology, Pancreas - metabolism, Pancreas - pathology, Pluripotency, Regenerative medicine, Reproductive Techniques, Assisted, RNA, Guide, CRISPR-Cas Systems - administration & dosage, Sequence Analysis, DNA, Sheep, Stem cell transplantation, Stem cells, Trans-Activators - genetics, Trans-Activators - metabolism