Abstract Growing switchgrass (Panicum virgatum, L.), a promising bioenergy crop, needs finely-tuned nitrogen (N) fertilization to improve biomass yields depending on soil types and site characteristics. N fertilization can also affect the soil organic carbon (SOC) pool. Therefore, this study was conducted to assess the effects of N fertilization on switchgrass biomass production and the SOC stock in Ohio. Switchgrass was established at three research stations (Northwest, Jackson, and Western sites) of the Ohio Agricultural Research and Development Center (OARDC) in spring 2004. N fertilizer was applied at four different rates (0, 50, 100, and 200 kg N ha−1) in 2008 and 2009. Aboveground and root biomass and the carbon (C) and N concentrations in plant tissues, SOC concentrations up to 30 cm depth were measured at the end of the growing season in 2009. Aboveground biomass at the Western site was the highest as 26 Mg ha−1 with 200 kg N ha−1 application, but there were no significant effects of N fertilization on aboveground biomass at two other sites and on root biomass across all sites. The amount of N export due to harvesting aboveground biomass increased with increase in N rates but did not vary among sites. With increasing N rates, the SOC stock linearly increased from 102 to 123 and from 55 to 70 Mg C ha−1 at the Northwest and the Jackson sites, respectively. However, this positive correlation was not observed for the Western site (a range of 59 to 67 Mg C ha−1). This study showed a potential of growing switchgrass as a bioenergy crop in Ohio and positive responses of the SOC stock to N fertilization.