Homologous recombination in human embryonic stem cells
- PMID: 12577066
- DOI: 10.1038/nbt788
Homologous recombination in human embryonic stem cells
Abstract
Homologous recombination applied to mouse embryonic stem (ES) cells has revolutionized the study of gene function in mammals. Although most often used to generate knockout mice, homologous recombination has also been applied in mouse ES cells allowed to differentiate in vitro. Homologous recombination is an essential technique if human ES cells are to fulfill their promise as a basic research tool. It also has important implications for ES cell-based transplantation and gene therapies. Significant differences between mouse and human ES cells have hampered the development of homologous recombination in human ES cells. High, stable transfection efficiencies in human ES cells have been difficult to achieve, and, in particular, electroporation protocols established for mouse ES cells work poorly in human ES cells. Also, in contrast to their murine counterparts, human ES cells cannot be cloned efficiently from single cells, making it difficult to screen for rare recombination events. Here we report an electroporation approach, based on the physical characteristics of human ES cells, that we used to successfully target HPRT1, the gene encoding hypoxanthine phosphoribosyltransferase-1 (HPRT1), and POU5F1, the gene encoding octamer-binding transcription factor 4 (Oct4; also known as POU domain, class 5, transcription factor 1 (POU5F1)).
Similar articles
-
Genetic recombination pathways and their application for genome modification of human embryonic stem cells.Exp Cell Res. 2010 Oct 1;316(16):2578-86. doi: 10.1016/j.yexcr.2010.06.004. Epub 2010 Jun 11. Exp Cell Res. 2010. PMID: 20542027 Review.
-
Oct4 expression revisited: potential pitfalls for data misinterpretation in stem cell research.Biol Chem. 2008 Jul;389(7):845-50. doi: 10.1515/BC.2008.098. Biol Chem. 2008. PMID: 18627312 Review.
-
Sall4 modulates embryonic stem cell pluripotency and early embryonic development by the transcriptional regulation of Pou5f1.Nat Cell Biol. 2006 Oct;8(10):1114-23. doi: 10.1038/ncb1481. Epub 2006 Sep 17. Nat Cell Biol. 2006. PMID: 16980957
-
Messenger RNA electroporation is highly efficient in mouse embryonic stem cells: successful FLPe- and Cre-mediated recombination.Gene Ther. 2004 Nov;11(21):1606-10. doi: 10.1038/sj.gt.3302342. Gene Ther. 2004. PMID: 15295620
-
Efficient gene targeting in mouse embryonic stem cells.Gene Ther. 1997 Jul;4(7):700-9. doi: 10.1038/sj.gt.3300457. Gene Ther. 1997. PMID: 9282171
Cited by
-
CRISPR-Cas12a-integrated transgenes in genomic safe harbors retain high expression in human hematopoietic iPSC-derived lineages and primary cells.iScience. 2023 Oct 27;26(12):108287. doi: 10.1016/j.isci.2023.108287. eCollection 2023 Dec 15. iScience. 2023. PMID: 38034357 Free PMC article.
-
A critical review on therapeutic approaches of CRISPR-Cas9 in diabetes mellitus.Naunyn Schmiedebergs Arch Pharmacol. 2023 Dec;396(12):3459-3481. doi: 10.1007/s00210-023-02631-1. Epub 2023 Jul 31. Naunyn Schmiedebergs Arch Pharmacol. 2023. PMID: 37522916 Review.
-
A Cell-Based Optimised Approach for Rapid and Efficient Gene Editing of Human Pluripotent Stem Cells.Int J Mol Sci. 2023 Jun 17;24(12):10266. doi: 10.3390/ijms241210266. Int J Mol Sci. 2023. PMID: 37373413 Free PMC article.
-
CRISPR/Cas9 and piggyBac Transposon-Based Conversion of a Pathogenic Biallelic TBCD Variant in a Patient-Derived iPSC Line Allows Correction of PEBAT-Related Endophenotypes.Int J Mol Sci. 2023 Apr 28;24(9):7988. doi: 10.3390/ijms24097988. Int J Mol Sci. 2023. PMID: 37175696 Free PMC article.
-
Transition from Animal-Based to Human Induced Pluripotent Stem Cells (iPSCs)-Based Models of Neurodevelopmental Disorders: Opportunities and Challenges.Cells. 2023 Feb 7;12(4):538. doi: 10.3390/cells12040538. Cells. 2023. PMID: 36831205 Free PMC article. Review.
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources
Medical
Research Materials
Miscellaneous