A comparative evaluation of three transfection procedures as assessed by resistance to G418 conferred to HEPG2 cells.
Three commonly used transfection techniques (electroporation, calcium phosphate precipitation and scrape loading) and a novel procedure combining the latter two methods were evaluated and conditions optimised for successful transfection of human HepG2 cells with plasmid DNA incorporating a mouse MHC Class I gene and a selectable marker (neomycin transferase gene) conferring resistance to G418. While transfection with linear DNA by scrape-loading gave satisfactory results, transfer of cloned circular DNA by electroporation, calcium phosphate precipitation or a combined use of scrape-loading and calcium phosphate gave best results for HepG2 cells.
Kumar
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S
Tenovus Laboratory, Faculty of Medicine, Southampton University Hospitals, England.
O'Dowd
C
C
Dunckley
M G
MG
Lund
T
T
D000818:Animals; D002130:Calcium Phosphates; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004270:DNA, Circular / Q000235:genetics; D004351:Drug Resistance; D018274:Electroporation; D005805:Genes, MHC Class I; D005839:Gentamicins / Q000494:pharmacology*; D006801:Humans; D019868:Kanamycin Kinase; D051379:Mice; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D010957:Plasmids; D014162:Transfection; D014407:Tumor Cells, Cultured
D002130:Calcium Phosphates; D004270:DNA, Circular; D005839:Gentamicins; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; C485838:calcium phosphate, monobasic, anhydrous; D004247:DNA; C020243:calcium phosphate; C010680:antibiotic G 418; D017853:Phosphotransferases (Alcohol Group Acceptor); D019868:Kanamycin Kinase; C485829:calcium phosphate, dibasic, anhydrous