Electric field-mediated gene transfer (electroporation) into mouse Friend and human K562 erythroleukemic cells.
Electroporation, the technique of electric field mediated gene transfer, was evaluated as a means of introducing and expressing genes into mouse Friend and human K562 erythroleukemic cells. Long-term (stable) gene expression in both Friend and K562 cells was measured using the recombinant plasmid Homer 6, which carries the aminoglycoside phosphotransferase (aph) gene as a selectable marker under the transcriptional control of the Moloney murine sarcoma virus long terminal repeat promoter/enhancer sequences. Parameters such as the DNA concentration, the initial field strength, the concentration of recipient cells, and the preselection expression time were examined to obtain optimal transfection frequencies. Short-term (transient) expression was also examined using the plasmid pLW4, which carries the chloramphenicol acetyltransferase gene under the transcriptional control of herpes simplex virus immediate early 5 gene promoter/enhancer sequences. Conditions that gave maximal stable transformation frequency were similar to those giving highest transient gene expression in the mouse and human erythroleukemic cell lines. Under optimal conditions, electroporation gave about ten times higher transfection frequencies and levels of transient expression for both types of cells when compared with the calcium phosphate technique. Because both Friend and K562 cells can be induced to differentiate in vitro, measurement of transient or stable expression levels for genes introduced into these cells may prove to be useful in the study of developmental regulation of genes from the erythroid pathway.
D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D004586:Electrophoresis / Q000379:methods; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000235:genetics; D007942:Leukemia, Experimental / Q000235:genetics*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics*; D051379:Mice; D010957:Plasmids; D014162:Transfection
D004273:DNA, Neoplasm; D015500:Chloramphenicol O-Acetyltransferase