Estimating soil nitrogen (N) mineralization is critical to balance fertilizer N requirements and their environmental impacts. In this study, net N mineralization was examined in soils under different crop rotations with each phase of the rotation present every year with biologically based incubations in 2011 and 2015. Net N mineralization was significantly different among treatments when the current crop was soybean, and the effect was dependent upon the previous crop and the cropping sequence. In particular, net increases in inorganic N were greater when the previous crop was winter wheat with or without red clover than if it were corn, and greater for the first year of soybean compared with the second year for rotations with two consecutive years of soybean in the 2011 incubation. However, cropping history did not influence net soil N mineralization when the current crop was corn, winter wheat, or winter wheat with red clover. In 2015, the presence of red clover in the rotation increased net N mineralization in all phases of the rotation. These results suggest both current and previous crops should be considered when estimating the N supplying capacity (net mineralization) of the soil. Net mineralizable N was found to be significantly correlated with total amino sugars (P < 0.001), glucosamine (P < 0.001), and galactosamine (P = 0.003), which suggests that amino sugars could be used as an indicator of the N supplying capacity of soil.