Aims <p>To explore the contribution of vertical root niche differentiation to recovery of deep NH₄⁺ by species in cultivated grass-clover mixtures, and to mixtures’ overyielding.</p> Methods <p>The uptake of a deep-placed (42&#xa0;cm) slow-release ¹⁵NH₄⁺ label was tracked in the herbage of two grass-clover mixtures during one growing season and the subsequent spring (3 + 1 harvests) and compared with uptake in monocultures.</p> Results <p>Mixtures overyielded in biomass (+ 11%), total N (+ 24%), and deep ¹⁵NH₄⁺ recovery (+ 17%). The latter was most pronounced in the autumn (+ 47% to + 62%). Diversity effects on deep ¹⁵N recovery were a combined result of changes in growth vigor and ¹⁵N recovery per biomass. We found evidence of vertical niche differentiation: <i>Schedonorus arundinaceus</i>, <i>Schedonorus pratensis</i> and <i>Lolium perenne</i> displayed positive diversity effects on deep ¹⁵N uptake per biomass combined with increased total N concentration; not so in <i>Phleum pratense</i> and <i>Poa pratensis</i>. Though the diversity effect on deep ¹⁵N per biomass was small in N-fixing <i>Trifolium pratense</i>, due to its high yields this translated into a large negative diversity effect on deep N recovery. Enhanced deep N uptake was contingent on enhanced growth vigor.</p> Conclusions <p>Improved N nutrition in grasses due to clovers did not prevent overall positive diversity effects on deep N recovery. This is potentially beneficial for reducing N leaching. The grass-clover mixtures better combined recovery of deep-placed ¹⁵NH₄⁺ with yield performance and high protein content than any single species. Diversity effects on deep N uptake were species-specific and could not be predicted by performance in monoculture.</p>