Recent studies have examined temporal fluctuations in the amount and carbon isotope content (δ13C) of CO2 produced by the respiration of roots and soil organisms. These changes have been correlated with diel cycles of environmental forcing (e.g., sunlight and soil temperature) and with synoptic-scale atmospheric motion (e.g., rain events and pressure-induced ventilation). We used an extensive suite of measurements to examine soil respiration over 2 months in a subalpine forest in Colorado, USA (the Niwot Ridge AmeriFlux forest). Observations included automated measurements of CO2 and δ13C of CO2 in the soil efflux, the soil gas profile, and forest air. There was strong diel variability in soil efflux but no diel change in the δ13C of the soil efflux (δR) or the CO2 produced by biological activity in the soil δJ). Following rain, soil efflux increased significantly, but δR and δJ did not change. Temporal variation in the δ13C of the soil efflux was unrelated to measured environmental variables, and we failed to find an explanation for this unexpected result. Measurements of the δ13C of the soil efflux with chambers agreed closely with independent observations of the isotopic composition of soil CO2 production derived from soil gas well measurements. Deeper in the soil profile and at the soil surface, results confirmed established theory regarding diffusive soil gas transport and isotopic fractionation. Deviation from best-fit diffusion model results at the shallower depths illuminated a pump-induced ventilation artifact that should be anticipated and avoided in future studies. There was no evidence of natural pressure-induced ventilation of the deep soil. However, higher variability in δ13C of the soil efflux relative to δ13C of production derived from soil profile measurements was likely caused by transient pressure-induced transport with small horizontal length scales.