( behavior. Two principally diff...)Transfection Reagent Selector Kit Handbook

behavior. Two principally different transfection techniques can be used; transient transfection and stable transfection. For further background information on transfection, please refer to current molecular biology manuals (2, 3).

Transient transfection

When cells are transiently transfected, the DNA is introduced into the nucleus of the cell, but does not integrate into the chromosome. This means that many copies of the gene of interest are present, leading to high levels of expressed protein. Transcription of the transfected gene can be analyzed within 2496 hours after introduction of the DNA depending on the construct used. Transient transfection is most efficient when supercoiled plasmid DNA is used.

Stable transfection

With stable or permanent transfection, the transfected DNA is either integrated into the chromosomal DNA or maintained as an episome. Although linear DNA results in lower DNA uptake by the cells relative to supercoiled DNA, it yields optimal integration of DNA into the host genome. Cells which have successfully integrated the DNA of interest or have maintained episomal plasmid DNA can be distinguished by using selectable markers. Frequently-used selectable markers are the genes encoding aminoglycoside phosphotransferase (APH; neoR gene) or hygromycin B phosphotransferase (HPH). Other selectable markers are the genes encoding adenosine deaminase (ADA), dihydrofolate reductase (DHFR), thymidine kinase (TK) or xanthine-guanine phosphoribosyl tranferase (XGPRT; gpt gene).

Primary Cells and Cell Lines

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