Optimization of electroporation parameters for the intramuscular delivery of plasmids in pigs.
Increased transgene expression after plasmid transfer to the skeletal muscle is obtained with electroporation in many species, but optimum conditions are not well defined. Using a plasmid with a muscle-specific secreted embryonic alkaline phosphatase (SEAP) gene, we have optimized the electroporation conditions in a large mammal (pig). Parameters tested included electric field intensity, number of pulses, lag time between plasmid injection and electroporation, and plasmid delivery volume. Electric pulses, between 0.4 and 0.6 Amp constant current, applied 80 sec after the injection of 0.5 mg SEAP-expressing plasmid in a total volume of 2 mL produced the highest levels of expression. Further testing demonstrated that electroporation of a nondelineated injection site reduces the levels of SEAP expression. These results demonstrate that electroporation parameters such as amperage, lag time, and the number of pulses are able to regulate the levels of reporter gene expression in pigs.
Khan
Amir S
AS
ADViSYS, Inc., The Woodlands, Texas 77381, USA.
Smith
Louis C
LC
Abruzzese
Ronald V
RV
Cummings
Kathleen K
KK
Pope
Melissa A
MA
Brown
Patricia A
PA
Draghia-Akli
Ruxandra
R
D000469:Alkaline Phosphatase / Q000097:blood / Q000235:genetics*; D000818:Animals; D018274:Electroporation / Q000379:methods*; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D018014:Gene Transfer Techniques; D007273:Injections, Intramuscular; D008297:Male; D018482:Muscle, Skeletal / Q000201:enzymology; D010957:Plasmids / Q000235:genetics*; D013552:Swine
D000469:Alkaline Phosphatase