Optimization of ectopic gene expression in skeletal muscle through DNA transfer by electroporation.
BACKGROUND
Electroporation (EP) is a widely used non-viral gene transfer method. We have attempted to develop an exact protocol to maximize DNA expression while minimizing tissue damage following EP of skeletal muscle in vivo. Specifically, we investigated the effects of varying injection techniques, electrode surface geometry, and plasmid mediums.


RESULTS
We found that as the amount of damage increased in skeletal muscle in response to EP, the level of beta-galactosidase (beta-gal) expression drastically decreased and that there was no evidence of beta-gal expression in damaged fibers. Two specific types of electrodes yielded the greatest amount of expression. We also discovered that DNA uptake in skeletal muscle following intra-arterial injection of DNA was significantly enhanced by EP. Finally, we found that DMSO and LipoFECTAMINE, common enhancers of DNA electroporation in vitro, had no positive effect on DNA electroporation in vivo.


CONCLUSIONS
When injecting DNA intramuscularly, a flat plate electrode without any plasmid enhancers is the best method to achieve high levels of gene expression.
Taylor
Jordan
J
Department of Basic Medical Sciences, Purdue University, W Lafayette, IN 47907, USA. jat4@cec.wustl.edu
Babbs
Charlie F
CF
Alzghoul
Mohammed Borhan
MB
Olsen
Aaron
A
Latour
Mickey
M
Pond
Amber L
AL
Hannon
Kevin
K
D000818:Animals; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D018274:Electroporation / Q000295:instrumentation / Q000379:methods*; D015870:Gene Expression / Q000187:drug effects; D007269:Injections, Intra-Arterial; D007273:Injections, Intramuscular; D007763:Lac Operon / Q000235:genetics; D008055:Lipids / Q000494:pharmacology; D051379:Mice; D018482:Muscle, Skeletal / Q000378:metabolism* / Q000473:pathology; D009336:Necrosis; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D014162:Transfection / Q000379:methods*; D001616:beta-Galactosidase / Q000378:metabolism
D008055:Lipids; C086724:Lipofectamine; D001616:beta-Galactosidase; D004121:Dimethyl Sulfoxide