Figure 3.
Creation of N. crassa strains expressing Tom40 variants. A sheltered heterokaryon containing a nucleus with a tom40RIP allele (Nucleus 1) and a nucleus with a wild-type tom40 allele (Nucleus 2) is depicted in the large box at the top of the figure. The heterokaryon was created by the procedure of sheltered RIP using strains described previously for linkage group V of N. crassa (Metzenberg and Grotelueschen, 1992). Additional markers in nucleus 1 impart cycloheximide resistance and requirements for lysine and leucine. Transformation of the strain with mutant alleles of tom40, carried on a bleomycin resistance plasmid, followed by selection on media containing bleomycin (3.7 μg/ml), cycloheximide (50 μg/ml), lysine, and leucine resulted in homokaryons expressing only the predicted mutant alleles of tom40.