(A) Multiple copies of TIM18
suppress the growth defect of tim54-1 mutants but not
tim23-1 mutants. tim54-1 his3 strain 735
was transformed with one of the following high-copy plasmids:
2μ-TIM18 plasmid pOK66, 2μ-TIM54
plasmid pOK59, 2μ-TIM22 plasmid pJH203, or the empty
vector pRS423 (Sikorski and Hieter, 1989). tim23-1 leu2
ura3 strain 574 was transformed with 2μ-TIM18
plasmid pOK66, 2μ-TIM23 plasmid pKR21 (Ryan et
al., 1998), 2μ-TIM17 plasmid pKR7 (Ryan
et al., 1994), or the empty vector pRS426 (Sikorski and
Hieter, 1989). Transformants were streaked onto YEPmedium containing 2% glycerol and ethanol
and incubated at 24 or 34°C for 5 d. (B) Multiple copies of
TIM18 do not suppress the
tim54::KAN or
tim22::TRP1 disruptions.
tim22::TRP1 strain 935, which carries the
TIM22-URA3 plasmid pJH202 (Kerscher et al., 1997), was
transformed with 2μ-TIM18 plasmid pOK66,
2μ-TIM22 plasmid pJH203, or the empty vector pRS423
(Sikorski and Hieter, 1989). tim54::KAN strain
1078, which carries the TIM54-URA3 plasmid pOK30
(Kerscher et al., 1997), was transformed with
2μ-TIM18 plasmid pOK66, CEN-TIM54
plasmid pOK22 (Kerscher et al., 1997), or the empty
vector pRS424 (Sikorski and Hieter, 1989). Transformants were patched
onto YEPD medium and then replica plated onto medium containing 5-FOA
to select for loss of the TIM22-URA3 or
TIM54-URA3 plasmids.