The Journal of Membrane Biology

, Volume 236, Issue 1, pp 81–85

Influence of Plasmid Concentration on DNA Electrotransfer In Vitro Using High-Voltage and Low-Voltage Pulses

  • Karolina Čepurnienė
  • Paulius Ruzgys
  • Rimantas Treinys
  • Ingrida Šatkauskienė
  • Saulius Šatkauskas
Article

DOI: 10.1007/s00232-010-9270-5

Cite this article as:
Čepurnienė, K., Ruzgys, P., Treinys, R. et al. J Membrane Biol (2010) 236: 81. doi:10.1007/s00232-010-9270-5

Abstract

DNA electrotransfer in vivo for gene therapy is a promising method. For further clinical developments, the efficiency of the method should be increased. It has been shown previously that high efficiency of gene electrotransfer in vivo can be achieved using high-voltage (HV) and low-voltage (LV) pulses. In this study we evaluated whether HV and LV pulses could be optimized in vitro for efficient DNA electrotransfer. Experiments were performed using Chinese hamster ovary (CHO) cells. To evaluate the efficiency of DNA electrotransfer, two different plasmids coding for GFP and luciferase were used. For DNA electrotransfer experiments 50 μl of CHO cell suspension containing 100, 10 or 1 μg/ml of the plasmid were placed between plate electrodes and subjected to various combinations of HV and LV pulses. The results showed that at 100 μg/ml plasmid concentration LV pulse delivered after HV pulse increased neither the percentage of transfected cells nor the total transfection efficiency (luciferase activity). The contribution of the LV pulse was evident only at reduced concentration (10 and 1 μg/ml) of the plasmid. In comparison to HV (1,200 V/cm, 100 μs) pulse, addition of LV (100 V/cm, 100 ms) pulse increased transfection efficiency severalfold at 10 μg/ml and fivefold at 1 μg/ml. At 10 μg/ml concentration of plasmid, application of four LV pulses after HV pulse increased transfection efficiency by almost 10-fold. Thus, these results show that contribution of electrophoretic forces to DNA electrotransfer can be investigated in vitro using HV and LV pulses.

Keywords

Gene transfer DNA electrotransfer DNA electrophoresis Gene expression Electroporation Electropermeabilization 

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Karolina Čepurnienė
    • 1
  • Paulius Ruzgys
    • 1
  • Rimantas Treinys
    • 1
    • 2
  • Ingrida Šatkauskienė
    • 1
  • Saulius Šatkauskas
    • 1
  1. 1.Biophysical Research Group, Biology DepartmentVytautas Magnus UniversityKaunasLithuania
  2. 2.Institute of CardiologyKaunas University of MedicineKaunasLithuania

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