DNA transfection. 1993

K Pardy
Neuropeptide Laboratory, Institute of Molecular and Cell Biology, National University of Singapore, Republic of Singapore.

Prior to the introduction of transgene constructs into animals, it is useful first to determine the presence of a functional promoter and transcription unit. This can be achieved by transient transfection of the construct into heterologous cell lines and subsequent measurement of reporter enzyme activity in cell extracts. Following ligation of the chosen promotor sequence to the reporter enzyme coding region in a suitable plasmid vector, DNA is purified and transfected into an appropriate cell line. Popular methods of transfection are the calcium phosphate method, electroporation, and lipofection. The latter method, although relatively expensive, provides a quick and reliable method of transfection and is the method described in this chapter. This method of transfection relies on a positively charged lipid, DOTMA (N [1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride), which forms liposomes that interact with DNA and RNA and carries them into mammalian cells in culture (1). DOTMA is available commercially and has been shown to be suitable for a variety of cell lines.

UI MeSH Term Description Entries
D002460 Cell Line Established cell cultures that have the potential to propagate indefinitely. Cell Lines,Line, Cell,Lines, Cell
D004247 DNA A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine). DNA, Double-Stranded,Deoxyribonucleic Acid,ds-DNA,DNA, Double Stranded,Double-Stranded DNA,ds DNA
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D014162 Transfection The uptake of naked or purified DNA by CELLS, usually meaning the process as it occurs in eukaryotic cells. It is analogous to bacterial transformation (TRANSFORMATION, BACTERIAL) and both are routinely employed in GENE TRANSFER TECHNIQUES. Transfections

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