Experiments of 13C and 15N Uptake Applied as Double-Labeled ATP in the Field

To test if ¹³C and ¹⁵N uptake rates calculated as ATP equivalents in experiments under controlled conditions were similar to ¹³C and ¹⁵N uptake rates equivalent to ATP in the field, ATP uptake experiments were performed in September 2017 at two field sites, namely the acidic Conventwald (Con) and calcareous Tuttlingen (Tut) forest stands. The soils of these forests differ in their properties (silicate versus limestone bedrock) (Prietzel et al., 2016) with the Tuttlingen soil containing eightfold lower plant available P_i (for detailed soil descriptions see Prietzel et al., 2016; Netzer et al., 2017). At both field sites, fine roots of six adult beech trees and of six beech saplings were carefully excavated out of the soil. Adherent soil particles from the roots were removed with distilled water and cleaned roots were dried using paper towels. Roots still attached to adult beech trees or to their offspring were incubated in an artificial soil solution at pH 5.0 that contained 29 μM NH₄Cl, 35 μM KNO₃, 16 μM CaCl₂, 17 μM MgCl₂ 0.3 μM MnCl₂, 22 μM NaCl, and 0.169 mM ATP. Double-labeled ATP was diluted to 10 atom% (ATP-¹³C₁₀¹⁵N₅, 98 atom%, Sigma Aldrich). Fine roots were cut from the trees after 4 h of incubation, rinsed with distilled water to remove adherent ATP-¹³C₁₀¹⁵N₅, dried in an oven (72 h, 50°C) for at least 2 days and homogenized using mortar and pestle.