Transfection by particle bombardment: delivery of plasmid DNA into mammalian cells using gene gun.
BACKGROUND
Recently, particle bombardment has become increasingly popular as a transfection method, because of a reduced dependency on target cell characteristics. In this study, we evaluated in vitro gene transfer by particle bombardment.


METHODS
gWIZ luciferase and gWIZ green fluorescent protein (GFP) plasmids were used as reporter genes. Mammalian cell lines HEK 293, MCF7 and NIH/3T3 were used in the transfection experiments. Transfection was performed by bombardment of the cells with gene-coated gold particles using the Helios Gene Gun. The technology was assessed by analyzing gene expression and cell damage. Cell damage was evaluated by MTT assay.


RESULTS
This technology resulted in efficient in vitro transfection, even in the cells which are difficult to transfect. The gene expression was dependent on the gene gun's helium pressure, the sizes of the gold particles, the amount of the particles and DNA loading, while cell viability was mostly dependent on helium pressure and amount of the gold particles.


CONCLUSIONS
This technology was useful to transfection of cells. Optimal transfection conditions were determined to be between 75 and 100 psi of helium pressure, 1.0 to 1.6 mum gold particle size and 0.5 mg of gold particle amount with a loading ratio of 4 microg DNA/mg gold particles.


CONCLUSIONS
These findings will be useful in the design of gene gun device, and bring further improvements to the in vitro and in vivo transfection studies including gene therapy and vaccination.
Uchida
Masaki
M
The School of Pharmacy, University of London, 29-39 Brunswick Square, London WC1N1AX, UK.
Li
Xiong Wei
XW
Mertens
Peter
P
Alpar
H Oya
HO
D000818:Animals; D019470:Biolistics / Q000379:methods*; D002460:Cell Line; D004247:DNA / Q000378:metabolism* / Q000648:ultrastructure; D004249:DNA Damage; D004587:Electrophoresis, Agar Gel; D005786:Gene Expression Regulation; D006046:Gold / Q000378:metabolism*; D006801:Humans; D008156:Luciferases / Q000235:genetics; D051379:Mice; D018613:Microscopy, Confocal; D010316:Particle Size; D010957:Plasmids / Q000378:metabolism* / Q000648:ultrastructure; D034741:RNA, Small Interfering / Q000378:metabolism; D014162:Transfection / Q000379:methods*
D034741:RNA, Small Interfering; D006046:Gold; D004247:DNA; D008156:Luciferases