A piggyBac-based platform for genome editing and clonal rhesus macaque iPSC line derivation
Autor: | Daniel Urrutia-Cabrera, Iris Bartels, Ralf Dressel, Anna Maierhofer, Yuliia Tereshchenko, Iga Grządzielewska, Ignacio Rodriguez-Polo, Felix Mattern, Thomas Haaf, Stoyan Petkov, Sophie Mißbach, Rüdiger Behr |
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Jazyk: | angličtina |
Rok vydání: | 2021 |
Předmět: |
0301 basic medicine
Male Science Genetic Vectors Green Fluorescent Proteins Induced Pluripotent Stem Cells Transposases Context (language use) Computational biology Biology Transfection Genome Article Stem-cell biotechnology Cell Line 03 medical and health sciences Mice 0302 clinical medicine Genome editing CRISPR-Associated Protein 9 CRISPR Animals Transgenes Induced pluripotent stem cell Phylogeny Skin Gene Editing Multidisciplinary Fibroblasts Cellular Reprogramming Embryonic stem cell Macaca mulatta humanities 030104 developmental biology PiggyBac Transposon System DNA Transposable Elements Medicine Female CRISPR-Cas Systems Reprogramming 030217 neurology & neurosurgery Biotechnology |
Zdroj: | Scientific Reports, Vol 11, Iss 1, Pp 1-17 (2021) Scientific Reports |
ISSN: | 2045-2322 |
Popis: | Non-human primates (NHPs) are, due to their close phylogenetic relationship to humans, excellent animal models to study clinically relevant mutations. However, the toolbox for the genetic modification of NHPs is less developed than those for other species like mice. Therefore, it is necessary to further develop and refine genome editing approaches in NHPs. NHP pluripotent stem cells (PSCs) share key molecular signatures with the early embryo, which is an important target for genomic modification. Therefore, PSCs are a valuable test system for the validation of embryonic genome editing approaches. In the present study, we made use of the versatility of the piggyBac transposon system for different purposes in the context of NHP stem cell technology and genome editing. These include (1) Robust reprogramming of rhesus macaque fibroblasts to induced pluripotent stem cells (iPSCs); (2) Culture of the iPSCs under feeder-free conditions even after removal of the transgene resulting in transgene-free iPSCs; (3) Development of a CRISPR/Cas-based work-flow to edit the genome of rhesus macaque PSCs with high efficiency; (4) Establishment of a novel protocol for the derivation of gene-edited monoclonal NHP-iPSC lines. These findings facilitate efficient testing of genome editing approaches in NHP-PSC before their in vivo application. |
Databáze: | OpenAIRE |
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