Temporal order of acquired mutations. DNA extracted from frozen intermediate time points of the adaptive evolutions was Sanger sequenced at genomic locations corresponding to mutations in the endpoints. Time points that were sequenced for mutations are indicated by an arrowhead. The arrow is white if no mutations were identified that were not identified at a previous time point. The first day each mutation was observed is indicated with a dark arrow. Curves represent the growth rate trajectory during the period of adaptive evolution. (a) LactA, (b) LactC, (c) LactD, (d) LactE. The atoS, acpP, and yjbM genes are not represented in the figure because they were not identified as penetrating more than 50% of the population by day 60 of adaptive evolution.