Electrophoretic component of electric pulses determines the efficacy of in vivo DNA electrotransfer.
Efficient DNA electrotransfer can be achieved with combinations of short high-voltage (HV) and long low voltage (LV) pulses that cover two effects of the pulses, namely, target cell electropermeabilization and DNA electrophoresis within the tissue. Because HV and LV can be delivered with a lag up to 3000 sec between them, we considered that it was possible to analyze separately the respective importance of the two types of effects of the electric fields on DNA electrotransfer efficiency. The tibialis cranialis muscles of C57BL/6 mice were injected with plasmid DNA encoding luciferase or green fluorescent protein and then exposed to various combinations of HV and LV pulses. DNA electrotransfer efficacy was determined by measuring luciferase activity in the treated muscles. We found that for effective DNA electrotransfer into skeletal muscles the HV pulse is prerequisite; however, its number and duration do not significantly affect electrotransfer efficacy. DNA electrotransfer efficacy is dependent mainly on the parameters of the LV pulse(s). We report that different LV number, LV individual duration, and LV strength can be used, provided the total duration and field strength result in convenient electrophoretic transport of DNA toward and/or across a permeabilized membrane.
Satkauskas
Saulius
S
Vectorology and Gene Transfer, UMR 8121 CNRS, Institute Gustave Roussy, F-94805 Villejuif, France.
André
Franck
F
Bureau
Michel F
MF
Scherman
Daniel
D
Miklavcic
Damijan
D
Mir
Lluis M
LM
D000818:Animals; D004247:DNA / Q000008:administration & dosage / Q000235:genetics; D018274:Electroporation / Q000379:methods; D005260:Female; D018014:Gene Transfer Techniques; D005819:Genetic Markers / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D018482:Muscle, Skeletal / Q000378:metabolism
D005819:Genetic Markers; D049452:Green Fluorescent Proteins; D004247:DNA; D008156:Luciferases