Microbial evolution is driven by environmental and ecological pressures and realized by selection for adaptive traits within affected populations. In recent decades, increased understanding of the abundance, diversity and activity of microbial inhabitants of marine subsurface sediments has illuminated ecological interactions and environmental variations in the deep biosphere. Upon this foundation, subsurface microbial evolutionary processes are ripe for exploration. To examine the interplay between phenotypic adjustment and evolutionary adaptation in the subsurface, and tradeoffs between metabolic capacity and growth in these energy-limited environments, a novel laboratory platform will be employed for the study of in vivo spatially resolved microbial experimental evolution. The platform is a large growth plate set up to mimic a subsurface sedimentary environment with sustained anoxia and progressive limitations in the abundance and quality of organic carbon. An isoclonal inoculum of sulfate reducing Desulfobacterium autotrophicum will be introduced at one end of the plate and monitored visually as the culture encounters and adapts to abrupt changes in organic carbon availability. Successful strains will be characterized for growth characteristics, adaptive mutations and isotopic fractionation. These experiments further C-DEBI Theme III and inform the tempo, mode and synonymous nature of microbial evolution for a substantial portion of Earth’s biomass.