We are characterizing a diploid strain of yeast that evolved under selection in a carbon-limited environment. This strain (L4) is being compared to the 'parent' strain (CP1AB) used to initiate a continuous culture run for 450 generations under glucose-limitation. The two strains differ in cell shape, colony size and growth on galactose. When grown over three days in glucose batch culture CP1AB shows typical stationary phase density changes; by contrast the density of viable L4 cells in 72 hr batch cultures is significantly lower than at 24 hrs, suggesting changes in L4's ability to respond appropriately to starvation. Steady-state density of L4 in chemostats was fourfold greater than CP1AB, suggesting that L4 is more efficient at either acquiring or utilizing glucose. This observation is consistent with steady-state glucose concentrations which are more than an order of magnitude lower for L4 than for CP1AB. In addition, transport assays on cells grown in chemostats demonstrate statistically greater activity and affinity for glucose among L4 cells. Restriction site analysis of HXT 4, a hexose transport gene that is highly expressed at low substrate concentrations reveals marked differences between L4 and CP1AB which suggest duplication of the HXT4 locus. RFLP analysis of other hexose transport genes and glycolysis genes reveal no such polymorphism. We are focusing our analysis of evolutionary change at the HXT4 and GAL2 loci on comparisons of mRNA levels.