Positive selection forDictyostelium mutants lacking uridine monophosphate synthase activity based on resistance to 5-fluoro-orotic acid

✍ Scribed by Kalpaxis, Dimitrios ;Werner, Herbert ;Marcotte, Emmanuell Boy ;Jacquet, Michel ;Dingermann, Theodor

• Publisher: John Wiley and Sons
• Year: 1990
• Tongue: English
• Weight: 674 KB
• Volume: 11
• Category: Article
• ISSN: 0192-253X
• DOI: 10.1002/dvg.1020110512

✦ Synopsis

Abstract

Whereas transformation of__Dictyostelium discoideum__ now can be done routinely and reliably, there is increasing demand for a system that allows selection of cells that have gone through repeated transformation cycles. Such a system is presented here. Selection is based on resistance to 5‐fluoro‐orotic‐acid (5‐FOA). D. discoi‐deum is highly sensitive to this drug, and cells can survive only in the presence of 5‐FOA if they carry a defective UMP‐synthase gene. After transformation with a plasmid carrying a cloned version of the UMP‐synthase gene, 5‐FOA resistant cells are obtained at high frequency. Because 5‐FOA‐resistant cells depend on exogenously added uracil, these cells can be taken through a second round of transformation. A plasmid‐borne UMP‐synthase gene renders 5‐FOA‐resistant cells phenotypically ura ^+^. Finally, cells can be further transformed using the standard G418 selection system.