Figure 2.

Tests to identify causative mutations in lineage 1. (A) Allele replacements were done to test each candidate mutation. Starting with polygenic mutant 1.2, three mutations mot3-1, grr1-1, and sgm1-1 were each replaced with the wild-type allele and were then tested for long-distance activation by expression of the HIS3 reporter on the plates shown. All plates contain galactose as the carbon source. Plates were incubated at 30° for 4 days. (B) To test the sufficiency of the identified causative mutations, beginning with the sin4Δ strain, FY3055, strains were constructed that contained each mutation singly (data not shown), as different combinations of double mutants, and as the mot3-1 grr1-1 sin4Δ0 triple mutant. All plates contain galactose as the carbon source. Plates were incubated at 30° for 4 days. (C) Northern analysis was done to measure HIS3 RNA levels in the reconstructed mutants and to compare the levels to that in the original polygenic mutant 1.2.