Figure 6.

Expression of a mitochondrial-targeted EcoRI endonuclease in yeast. (A) The effect of EcoRI expression on growth in glycerol. Cultures were grown to mid-log phase in glucose. Yeast were sedimented by centrifugation, incubated for 30 min at the non-permissive temperature in galactose and their growth rate in glycerol subsequently determined. (Left) Wild-type incubated in galactose (filled squares); wild-type harboring the CDC9–EcoRI fusion gene not induced with galactose (filled triangles); wild-type harboring the CDC9–EcoRI gene induced with galactose (filled circles). (Right) cdc9-1^ts incubated in galactose (open squares); cdc9-1^ts harboring the CDC9–EcoRI fusion gene not induced with galactose (open triangles); cdc9-1^ts harboring the CDC9–EcoRI fusion gene induced with galactose (open circles). (B) Analysis of mitochondrial DNA. Genomic DNA was isolated from yeast clones treated as described above (except that the incubation in galactose was performed at the non-permissive temperature; see text) and mitochondrial DNA analyzed. (Upper) Southern blot hybridization; (lower) ethidium bromide stained agarose gel. Wild-type yeast (WT), wild-type yeast harboring the CDC9–EcoRI fusion gene (WT-RI), cdc9-1^ts yeast and cdc9-1^ts yeast harboring the CDC9–EcoRI fusion gene (cdc9-1^ts-RI) were examined. Three conditions were analyzed: yeast not exposed to galactose (–); yeast exposed to galactose (+); yeast exposed to galactose and permitted to recover in glycerol medium (R).