In the southern United States, much of the emphasis in bottomland restoration is placed on establishing an oak-dominated forest. Artificial regeneration is an alternative for restoration on cleared lands and where a desirable seed source is not present. Currently the standard procedure for seedling preparation is to prune the roots prior to transplanting in the field. It is not fully known what effect(s) root pruning has on transplanted seedlings. In addition, bottomland restoration efforts inherently take place on floodplains. The potential interaction between root pruning and flooding on seedling performance is not known. This study consisted of two separate but related laboratory experiments. The purpose of the first experiment was to quantify the effects of various percentages of root removal and varying soil moisture regimes on transplanted Nuttall oak seedlings (Quercus nuttallii Palmer). Root pruning treatments consisted of removal of roots at 0%, 25% and 75% while soil moisture regime was maintained at non-flooded or periodically flooded conditions. Plant gas exchange, growth, and survival were measured. Root pruning alone had adverse effects on height growth during the first 72 days following transplanting. Periodic flooding also produced adverse effects on stomatal conductance (p = 0.0002), height growth (p = 0.005), and survival (p = 0.02). Photosynthetic data indicated that as pruning intensified in the periodically flooded seedlings, photosynthetic rates decreased. In contrast, as pruning intensified in the non-flooded seedlings, photosynthesis increased. This demonstrated that pruning rate had a varying effect on photosynthesis dependent upon soil moisture condition. Experiment 2 focused on the effects of varying degrees of root pruning on new root formation. The seedlings were grown under laboratory conditions, harvested at 0, 10, 20, and 30 days after treatment initiation, and analyzed for new root formation. Results of Experiment 2 indicated no difference in new root formation, root length, or root biomass due to the pruning treatment. Overall, our results from both experiments indicated that root pruning had no detectable long-term adverse effects on growth and survival of seedlings under drained soil conditions; however, as results from Experiment 1 demonstrated, if seedlings were planted in periodically flooded conditions, root pruning produced adverse effects. Thus, in restoration efforts utilizing Nuttall oak seedlings, the planting strategy and pruning rate should be carefully evaluated based on the knowledge of sites' hydrology. Alternatively, on sites with unpredictable flooding both pruned and unpruned seedlings may be utilized to ensure survival.