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Escherichia coli lab strains K-12 GM4792 Lac and GM4792 Lac carry opposite lactose markers, which are useful for distinguishing evolved lines as they produce different colored colonies. The two closely related strains are chosen as ancestors for our ongoing studies of experimental evolution. Here, we describe the genome sequences, annotation, and features of GM4792 Lac and GM4792 Lac. GM4792 Lac has a 4,622,342-bp long chromosome with 4,061 protein-coding genes and 83 RNA genes. Similarly, the genome of GM4792 Lac consists of a 4,621,656-bp chromosome containing 4,043 protein-coding genes and 74 RNA genes. Genome comparison analysis reveals that the differences between GM4792 Lac and GM4792 Lac are minimal and limited to only the targeted lac region. Moreover, a previous study on competitive experimentation indicates the two strains are identical or nearly identical in survivability except for lactose utilization in a nitrogen-limited environment. Therefore, at both a genetic and a phenotypic level, GM4792 Lac and GM4792 Lac, with opposite neutral markers, are ideal systems for future experimental evolution studies. [ABSTRACT FROM AUTHOR]