Recently, root traits have been suggested to play an important role in developing greater nitrogen uptake and grain yield. However, relatively few breeding programs utilize these root traits. Over a series of experiments at different growth stages with destructive plant biomass measurements, we analyzed above-ground and below-ground traits in seven geographically diverse lines of wheat. Root and shoot biomass allocation in 14-day-old seedlings were analyzed using paper roll-supported hydroponic culture in two Hoagland solutions containing 0.5 (low) and 4 (high) mM of nitrogen (N). For biomass analysis of plants at maturity, plants were grown in 7.5 L pots filled with soil mix under two nitrogen treatments. Traits were measured as plants reached maturity. High correlations were observed among duration of vegetative growth, tiller number, shoot dry matter, and root dry matter. Functionality of large roots in nitrogen uptake was dependent on the availability of N. Under high N, lines with larger roots had a greater yield response to the increase in N input. Under low N, yields were independent of root size and dry matter, meaning that there was not a negative tradeoff to the allocation of more resources to roots, though small rooted lines were more competitive with regards to grain yield and grain N concentration in the low-N treatment. In the high-N treatment, the large-rooted lines were correlated to an increase in grain N concentration (r = 0.54) and grain yield (r = 0.43). In low N, the correlation between root dry matter to yield (r = 0.20) and grain N concentration (r = −0.38) decreased. A 15-fold change was observed between lines for root dry matter; however, only a ~5-fold change was observed in shoot dry matter. Additionally, root dry matter measured at the seedling stage did not correlate to the corresponding trait at maturity. As such, in a third assay, below-ground and above-ground traits were measured at key growth stages including the four-leaf stage, stem elongation, heading, post-anthesis, and maturity. We found that root growth appears to be stagnant from stem elongation to maturity.