The origin and fate of genetic diversity are central organizing themes in biology, crucial to discovering how life arose and evolved on earth, and how life might arise and evolve elsewhere in the universe. Especially relevant is how diversity originates and is molded in the absence of sexual recombination and in the presence of one or few limiting resources. Classical models of evolution lead to the expectation that under these conditions population genetic diversity should be limited in scope and transient in nature, as selection favors successively fitter clones. However, empirical and theoretical evidence now indicate otherwise. Even under simple laboratory conditions, asexual populations can become highly diverse owing to clonal interference, where beneficial mutations arise in multiple competing lineages. Our group is exploring another mechanism by which diversity can increase in such populations, one where mutations arising in different lineages result in cooperative resource use via mutually beneficial interactions, which we call clonal reinforcement. We have carried out long-term evolution experiments using Escherichia coli cultured under continuous glucose limitation, and identified ancestral mutations in intermediary metabolism that may favor this evolutionary trajectory. Phylogenetic reconstruction reveals that lineages giving rise to such simple communities arise early and diverge rapidly. We also discovered mutations unique to the dominant community member that appear to enhance glucose scavenging and maintain redox balance, but do so at the expense of carbon lost to overflow metabolites. Because these metabolites serve as growth substrates that are differentially accessible to the other community members, we conclude that this lineage likely serves as an “engine” generating diversity by creating new metabolic niches, but not the occupants themselves. Laboratory experimental evolution therefore has the power to illuminate the emergence of syntrophy, a ubiquitous feature of microbial communities in nature including those that cause chronic infection.