Bacterial IMPDH gene used for the selection of mammalian cell transfectants.
Stable cell transfection is used for the expression of exogenous genes or cDNAs in eukaryotic cells. Selection of these transfectants requires a dominant selectable marker. A variety of such markers has been identified and is currently in use. However, many of these are not suitable for all cell types or require unique conditions. Here we describe a simple and versatile dominant selectable marker that involves bacterial IMP dehydrogenase (IMPDH), an enzyme essential for the replication of mammalian and bacterial cells. Although IMPDH is evolutionarily conserved, the bacterial enzyme is orders of magnitude more resistant to the toxic effect of the drug mycophenolic acid, which is an IMPDH inhibitor. We have demonstrated that transfection of human, monkey or Chinese hamster cell lines with an expression vector containing bacterial IMPDH and mycophenolic acid treatment resulted in the selection of colonies with a strikingly increased resistance to mycophenolic acid toxicity. Analysis of cells derived from these colonies indicated that the acquisition of this resistance was associated with bacterial IMPDH protein expression. As a proof of principle, we showed that mammalian cell transfection with a hicistronic IMPDH/GFP expression vector and mycophenolic acid treatment can he used to successfully select transfectants that express the fluorescent protein. These results indicate that bacterial IMPDH is a practical dominant selectable marker that can be used for the selection of transfectants that express exogenous genes or cDNAs in mammalian cells.
- Research Organization:
- Argonne National Lab. (ANL), Argonne, IL (United States)
- Sponsoring Organization:
- National Institutes of Health (NIH)
- DOE Contract Number:
- DE-AC02-06CH11357
- OSTI ID:
- 961223
- Report Number(s):
- ANL/ES/JA-45540; TRN: US201011%%496
- Journal Information:
- BioTechiques, Vol. 34, Issue 6 ; Jun. 2003; ISSN 0736-6205
- Country of Publication:
- United States
- Language:
- ENGLISH
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