•18 sites of field experiments were conducted in two provinces of China for testing the relation of wheat responses to N fertilizer with soil accumulated nitrate N in 0–100cm depth.•Soil ammonium and nitrate N were measured from 0 to 100cm and mineralized N during wheat growing period estimated.•On average, in 0–100cm depth, soil accumulated 132kg nitrate N and 150.8kg of ammonium and nitrate N.•Cumulative nitrate N was closely correlated with control yield and yield increase by N addition. Eleven field trials of wheat (Triticum aestivum) in Yongshou County, Shaanxi Province and seven in Luoyang District, Henan Province were conducted with two treatments (without N as control and with 150kg Nha−1 addition). Soil nitrate N and ammonium N concentrations were determined in five layers from 0 to 100cm depth at each site. Results showed that soil ammonium concentrations were very low and concentrations and cumulative amounts had no relations with wheat yield and yield increase by N applications. Nitrate N concentrations, in contrast, were high and correlated with seed (grain) and biomass yields of wheat without added N, and with yield increase by N addition in some layers. The cumulative nitrate N in 0–20cm layer plus other layers to different depths was significantly or very significantly correlated with control yield and yield increase by N addition. The cumulative nitrate N in 0–80cm depth explained 70% of seed yield, and that in 0–100cm 72% shoot variation and over 55% yield increases. Nitrate N occupied 88% of the total amount of ammonium N and nitrate N, and had the same trends as the total in reflecting soil N supplying capacity. Wheat mainly depended on 0–20cm layer nitrate N at over-wintering stage, 0-40cm at reviving, 0-60cm or deeper at elongation stages, and at maturity, the correlation of cumulative nitrate N in 0–100cm layer was highest with biomass and biomass increase by N addition. The control wheat in Yongshou County absorbed 107.4kg Nha−1 from soil on average, equivalent to 72% of the cumulative nitrate N or 65% of the total mineral N in 0–100cm layer. After wheat harvest, the concentration of ammonium N was not significantly different from the initial while that of nitrate N greatly decreased.