Direct selection of stabilised yeast URA3 transformants with 5-fluorouracil

Yeast expression vectors are generally based on the 2 micron plasmid and are not stably inherited, so that continued selection pressure must be applied to prevent the gradual accumulation of plasmid-free cells. A number of 'autoselection ' systems have been proposed and perhaps the most versatile is the stabilisation of plasmids containing the URA3 gene in urc3 furl host strains (1). These strains are non-viable since they are blocked in uridine 5'-monophosphate synthesis by both the de novo and salvage pathways; maintenance of a WJ/13-containing plasmid is then obligatory for viability. Since the ura3 furl host strain is non-viable, the final transformant has been obtained by the lengthy procedure of mating a furl strain with a ura3 strain containing the URA3 plasmid, and selecting plasmid-containing ura3 furl progeny (1). It has been suggested (1) that spontaneous furl mutants of a transformant could be directly selected in one ste