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Fig. 4. Complementation of S. cerevisiae rho1 mutants by AgRHO1 genes. (A) Complementation of Scrho1. The open reading frame of ScRHO1 was replaced by either AgRHO1a or AgRHO1b in a diploid strain. After sporulation, the resulting strains were subjected to tetrad analysis. Spore growth is shown for AgRHO1a (left) and AgRHO1b (right). (B) Plasmid loss. Strains were transformed with RHO1L, a plasmid containing the URA3 marker and the ScRHO1 gene with promoter and terminator, and streaked on medium without or with counterselection for URA3 by 5'-fluorootic acid. (C) Two segregants from a tetrad shown in A (right panel) were tested for temperature sensitivity at 39°C in a drop dilution assay. Cultures were grown to an OD₆₀₀ of 1, the number of cells given below the image was spotted on agar plates and incubated for 3 days. (D) Complementation of Scrho1 temperature-sensitive mutants from different complementation groups by the two AgRHO1 genes. Strains carrying the wild-type ScRHO1 allele or an allele of each Scrho1 complementation group were combined with a Scrho1 deletion (top row), AgRHO1a (middle row) or AgRHO1b (bottom row). A drop dilution assay of cultures grown to OD₆₀₀=1 was performed. The strains were incubated at the permissive (25°C, left) or the restrictive temperature (39°C, right).

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