Electroporation optimization to deliver plasmid DNA into dental follicle cells

Yao, Shaomian; Rana, Samir Kumar; Liu, Dawen; Wise, Gary E.

doi:10.1002/biot.200900039

Cited by 28 publications

(21 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We noticed that with increased plasmid concentrations, the transfection efficiency is increased, however at the same time the cell viability is reduced. This finding is in agreement with a previous study [33]. Here, the optimal concentration of plasmid DNA for the transfection of BFFs was determined to be 10 μg per 100 μl.…”

Section: Discussionsupporting

confidence: 92%

Systematic optimization of square-wave electroporation conditions for bovine primary fibroblasts

Hyder

Eghbalsaied

2020

BMC Mol and Cell Biol

View full text Add to dashboard Cite

Background: Gene transfer by electroporation is an established method for the non-viral mediated transfection of mammalian cells. Primary cells pose a particular challenge for electroporation-mediated gene transfer, since they are more vulnerable than immortalized cells, and have a limited proliferative capacity. Improving the gene transfer by using square wave electroporation in difficult to transfect cells, like bovine fetal fibroblasts, is a prerequisite for transgenic and further downstream experiments. Results: Here, bovine fetal fibroblasts were used for square-wave electroporation experiments in which the following parameters were systematically tested: electroporation buffer, electroporation temperature, pulse voltage, pulse duration, pulse number, cuvette type and plasmid DNA amount. For the experiments a commercially available square-wave generator was applied. Post electroporation, the bovine fetal fibroblasts were observed after 24 h for viability and reporter expression. The best results were obtained with a single 10 millisecond square-wave pulse of 400 V using 10 μg supercoiled plasmid DNA and 0.3 × 10 6 cells in 100 μl of Opti-MEM medium in 4 mm cuvettes. Importantly, the electroporation at room temperature was considerably better than with pre-cooled conditions. Conclusions: The optimized electroporation conditions will be relevant for gene transfer experiments in bovine fetal fibroblasts to obtain genetically engineered donor cells for somatic cell nuclear transfer and for reprogramming experiments in this species.

show abstract

Section: Discussionsupporting

confidence: 92%

Systematic optimization of square-wave electroporation conditions for bovine primary fibroblasts

Hyder

Eghbalsaied

2020

BMC Mol and Cell Biol

View full text Add to dashboard Cite

show abstract

“…100 µM instead of 10 µM) fluorescence was detectable in small CD34+ cells. However, an undue concentration of nucleic acid can affect the cellular viability .…”

Section: Results and Discussionmentioning

confidence: 99%

Biophysical effects in off-resonant gold nanoparticle mediated (GNOME) laser transfection of cell lines, primary- and stem cells using fs laser pulses

et al. 2014

View full text Add to dashboard Cite

Gold nanoparticle mediated (GNOME) laser transfection is a powerful technique to deliver small biologically relevant molecules into cells. However, the transfection of larger and especially negatively charged DNA remains challenging. The efficiency for pDNA was 0.57% using parameter that does not influence the endo- and exogenous DNA. In order to gain a deeper understanding of the actual molecule uptake process, the uptake efficiency was determined using molecules of different sizes. It was evaluated that uncharged dextran molecules (2000 kDa) were delivered with an efficiency of 68%. The intracellular distribution of injected molecules was visualized and larger molecules were primary found in the cytoplasm. Patch clamp measurements suggested a permeabilization time up to 15 minutes. The uptake efficiency depended on the size and charge of the molecule to deliver as well as the cell size. A minor role for transfection plays the cell type since primary stem cells were successfully transfected. The perforation efficiency of semi-adherent and suspension cells is influenced by the cell and molecule size.

show abstract

“…EP has been incorporated for drug and gene delivery in various areas of the human body that include lung [88–90], skin [91–93], cornea [94, 95], spinal cord [96, 97], heart [98, 99], brain [100–102] and dental tissues [103, 104]. Owing to its high gene transfection efficiency, safety, low toxicity and reproducibility, EP also has the potential for introducing genes into mammalian cells [105].…”

Section: Electric Field Drug and Gene Delivery: Electroporation And Imentioning

confidence: 99%

Physical energy for drug delivery; poration, concentration and activation

Lakshmanan¹,

Gupta²,

Avci³

et al. 2014

Advanced Drug Delivery Reviews

156

103

View full text Add to dashboard Cite

Electroporation optimization to deliver plasmid DNA into dental follicle cells

Cited by 28 publications

References 27 publications

Systematic optimization of square-wave electroporation conditions for bovine primary fibroblasts

Systematic optimization of square-wave electroporation conditions for bovine primary fibroblasts

Biophysical effects in off-resonant gold nanoparticle mediated (GNOME) laser transfection of cell lines, primary- and stem cells using fs laser pulses

Physical energy for drug delivery; poration, concentration and activation

Contact Info

Product

Resources

About