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Plasmid DNA size does affect nonviral gene delivery efficiency in stem cells.
- Source :
-
Cellular reprogramming [Cell Reprogram] 2012 Apr; Vol. 14 (2), pp. 130-7. Date of Electronic Publication: 2012 Feb 17. - Publication Year :
- 2012
-
Abstract
- Genetic modification of stem cells, prior to transplantation, can enhance their survival and can improve their function in cell therapy settings. Mesenchymal stem cells (MSC) are considered one of the most promising tools for cell-based gene therapy, due to their multipotency, ease of isolation, as well as their high ex vivo expansion potential. Neural stem cells (NSC) may also present an ideal route for gene therapy and have been considered for use in cell replacement therapies in various neurodegenerative diseases. Gene therapy-based applications require the transfer of genetic material, either by viral or nonviral gene delivery methods, although the latter has been associated with low efficiencies, especially within hard to transfect cells as stem cells. Herein, we present results on the influence of plasmid size in gene delivery to human MSC and mouse NSC. We used minimized plasmids encoding a fluorescent protein but lacking the antibiotic resistance gene. This work shows that (1) for smaller plasmids the intracellular plasmid copy number can be up to 2.6-fold higher, and (2) the number of cells presenting fluorescence can be twice the number obtained for larger plasmids. Furthermore, by using plasmid constructs containing different polyA signals, we also demonstrated that differences between the plasmids depend largely on the transgene mRNA level. Based on our data we demonstrate that plasmid size severely affects the efficiency of nuclear uptake and we propose that it can also affect the rate of heterochromatin associated gene silencing in stem cells.
- Subjects :
- Animals
Cells, Cultured
DNA genetics
Efficiency
Electroporation
Genetic Vectors genetics
Green Fluorescent Proteins genetics
Humans
Mice
Stem Cells cytology
Stem Cells physiology
Transfection methods
Base Sequence physiology
Gene Transfer Techniques
Plasmids genetics
Stem Cells metabolism
Transgenes genetics
Subjects
Details
- Language :
- English
- ISSN :
- 2152-4998
- Volume :
- 14
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Cellular reprogramming
- Publication Type :
- Academic Journal
- Accession number :
- 22339198
- Full Text :
- https://doi.org/10.1089/cell.2011.0093