Determining Root and Fungal Derived Soil Organic Carbon Accumulate Rates v1

This protocol quantifies net incorporation of new belowground carbon (C) into a defined soil substrate using the δ¹³C contrast between C3 vegetation inputs and a C4-derived soil fill. The design partitions root + hyphae inputs from hyphae-only inputs using a nested mesh insert, and uses paired no-ingrowth controls to correct for plot-specific background shifts and water-mediated (dissolved organic carbon) inputs. This method can be used to estimate rates of soil organic matter formation from belowground sources; however, it does not reflect long-term stability and persistence of SOM. Scope and principle Isotopic tracer: C4 soil (relatively enriched δ¹³C) packed into cores placed under C3 vegetation (relatively depleted δ¹³C). Biological partitioning: A hyphae-only insert (37 µm mesh) nested within a root + hyphae outer core (3 mm mesh) separates hyphal-only from combined root + hyphal inputs. Control correction: A paired no-ingrowth control (1 µm mesh wrap) corrects for non-ingrowth isotopic shifts (e.g., plot specific background changes, water movement, DOC inputs).