Dominance of warm-season grasses modulates tallgrass prairie ecosystem structure and function. Reintroduction of these grasses is a widespread practice to conserve soil and restore prairie ecosystems degraded from human land use changes. Seed sources for reintroduction of dominant prairie grass species include local (non-cultivar) and selected (cultivar) populations. The primary objective of this study was to quantify whether intraspecific variation in developing root systems exists between population sources (non-cultivar and cultivar) of two dominant grasses (Sorghastrum nutans and Schizachyrium scoparium) widely used in restoration. Non-cultivar and cultivar grass seedlings of both species were isolated in an experimental prairie restoration at the Konza Prairie Biological Station. We measured above- and belowground net primary production (ANPP and BNPP, respectively), root architecture, and root tissue quality, as well as soil moisture and plant available inorganic nitrogen (N) in soil associated with each species and source at the end of the first growing season. Cultivars had greater root length, surface area, and volume than non-cultivars. Available inorganic N and soil moisture were present in lower amounts in soil proximal to roots of cultivars than non-cultivars. Additionally, soil NO3–N was negatively correlated with root volume in S. nutans cultivars. While cultivars had greater BNPP than non-cultivars, this was not reflected aboveground root structure, as ANPP was similar between cultivars and non-cultivars. Intraspecific variation in belowground root structure and function exists between cultivar and non-cultivar sources of the dominant prairie grasses during initial reestablishment of tallgrass prairie. Population source selection should be considered in setting restoration goals and objectives.