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The response of above- and below-ground biomass to soil nutrient availability is crucial for estimating below-ground carbon input and predicting changes in soil carbon storage. However, the response is far from clear at plant community level, especially for grassland systems. Using a long-term field experiment initiated 123 years ago with varying soil nutrient levels (deficient, sub-optimal, optimal and over-optimal) established by use of two nutrient sources (animal manure or mineral fertiliser), we examined the effects of soil nutrient level and source on herbage yield and composition, root biomass and root-to-shoot (R/S) ratio of an unfertilised multispecies grass-legume ley. Increased nutrient levels enhanced herbage yield, but did not affect root biomass. The R/S ratio decreased from deficient to sub-optimal level, but remained constant from optimal to over-optimal level. Nutrient source did not influence herbage yield, root biomass or R/S ratio, but the legume proportion increased in soils previously receiving mineral fertiliser. The R/S ratio decreased with herbage yield, but did not vary with herbage composition. We conclude that soil nutrient level and herbage yield rather than nutrient source and herbage composition determine biomass allocation between aboveground and belowground in temperate grassland leys.