The current study demonstrated that increasing N supply stimulated wheat root development by improving root morphological traits (RLD, RSA and DW) under field conditions. There were significant and positive correlations between these root morphological traits and root Zn uptake. These results indicated that the positive effects of N supply on root development were a contributing factor for improving root Zn uptake during all growth stages. To our knowledge, this is the first study to examine the relationships between root morphological traits and Zn uptake at different N application rates under field conditions. Furthermore, the greater ratios of shoot to root Zn concentration and shoot to total Zn content with increasing N supply suggests that optimising N supply promotes the root-to-shoot Zn translocation and results in greater overall shoot Zn accumulation. Shoot Zn accumulation positively correlated with grain yield and grain Zn concentration. This relationship indicates that higher shoot Zn accumulation, resulting from greater N supply, may maximize grain yield and grain Zn concentration. The positive effects of N on root Zn uptake, root-to-shoot Zn translocation, and Zn accumulation in grain and shoot are of significance for Zn biofortification strategies in cereals and improving grain yields.