Polybrene-mediated transfection of cultured lepidopteran cells: induction of a Drosophila heat shock promoter.
We have introduced hsp-cat plasmid DNA into Spodoptera frugiperda (Lepidoptera: Noctuidae) cells by transfection with purified DNA (1 to 48 micrograms/ml) mixed with the polycation polybrene (100 micrograms/ml) in serum-free Grace's medium. The hsp-cat construct contains a gene coding for the bacterial enzyme chloramphenicol acetyltransferase (CAT), whose expression is controlled by a promoter derived from a Drosophila heat shock (hsp) gene. Expression of CAT activity in transfected Spodoptera cells was induced by a 2-h heat shock at 43 degrees C. The temperature of the heat shock was based on conditions that maximized the expression of endogenous heat shock protein genes in these cells. CAT activity was maximal in cells that were exposed to the heat shock 2 d after transfection; by 4 d, activity was diminished, and little activity was detectable after 6 d. Transfection frequencies, which varied with DNA concentration and ranged as high as 6000 per million cells, were determined using a histochemical staining procedure.
Helgen
J C
JC
Department of Entomology, University of Minnesota, St. Paul 55108.
Fallon
A M
AM
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004121:Dimethyl Sulfoxide; D004330:Drosophila / Q000235:genetics; D006360:Heat-Shock Proteins / Q000235:genetics*; D006583:Hexadimethrine Bromide; D006358:Hot Temperature; D007915:Lepidoptera / Q000235:genetics*; D009036:Moths / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D014176:Protein Biosynthesis / Q000502:physiology; D013696:Temperature; D014162:Transfection
D006360:Heat-Shock Proteins; D006583:Hexadimethrine Bromide; D015500:Chloramphenicol O-Acetyltransferase; D004121:Dimethyl Sulfoxide