Rescue of transfected genes from mammalian cells by functional selection in Escherichia coli.
We have established procedures for reisolating a transfected gene from mammalian cells by selection in Escherichia coli for the function of the gene product using the Herpes simplex virus thymidine kinase gene as a model. Rescue of the gene is accomplished by three different methods. The tk gene is recloned into a plasmid in which it is hooked up by either the lac promoter or a lac/tk hybrid promoter, or the original plasmid is cut out of the host cell DNA. As the lac/tk hybrid gene can be expressed and selected both in the mammalian and E. coli cells, this type of gene rescue allows investigations on mutagenesis and methylation processes. Additionally, it offers a simple way of studying the integration of the transfected gene into the mammalian genome.
D000818:Animals; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005814:Genes, Viral; D007739:L Cells / Q000201:enzymology; D051379:Mice; D010957:Plasmids; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics*; D013937:Thymidine Kinase / Q000235:genetics*; D014162:Transfection
D013937:Thymidine Kinase; D004262:DNA Restriction Enzymes