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Development of tetraploid simplex strains.
Within Diploids 1 and 2, which have opposite mating types, one copy of the gene of interest (MFG 1, i.e., my favorite gene) was inactivated in each diploid by transformation with kanamycin or hygromicin cassettes that target deletions. The resulting diploids were crossed to create Met+ (see Materials and Methods) KanR and HygR tetraploids with two copies (duplex) of MFG1. Finally, a third copy of MFG1 was inactivated by transforming the duplex strain with a URA3 targeting cassette.