Figure 2

MSE sequence variations in vivo affect RNA levels through the SPO77 promoter. Shown at the bottom is a schematic of the heterologous in vivo chromosome constructs at the SPO77 locus. At one SPO77 locus, we replaced SPO77 with GFP, and the wild type MSE with mse*, which differs from the wild type by a single base mutation, written in capital letters (Except for (a), where the mse* is the same as the wild type.). The second SPO77 locus is unaltered. The resulting diploid SK1 strains were sporulated for 24 hours in liquid. Whole genomic RNA was extracted from these cells at 0,2,4,6,8,10, and 12 hours of sporulation, run on a denaturing polyacrylamide gel, transferred to a nitrocellulose membrane and probed sequentially with ³²P labeled probes to SPO77, GFP and PFY1. Included in the parenthesis is the in vitro binding of Ndt80 to each MSE variant relative to the wild type.