Tropical forests are important contributors for N2O emissions from soil to the atmosphere; however, the microbial processes responsible for soil N2O production in tropical forest soils remain unclear. In this study, we employed a 40-day-long in situ (NO3-)-N-15 labeling experiment to determine the microbial processes that are responsible for N2O production in an undisturbed tropical rainforest in Xishuangbanna, southwest China. We observed that during the experiment, most of the N-15-labeled NO3- were immobilized and only 0.027% was emitted as N2O. Of the total N2O production, 3.1% was from the added N-15-labeled NO3-, 48.6% was from the added NH4+ and native N. Within the first 27 h, nitrification (heterotrophic and autotrophic nitrification) was the predominant microbial process of N2O production, accounting for 97.6% of total soil N2O emissions, whereas denitrification and co-denitrification resulted in only 1.6% and 0.9% emissions, respectively. To the best of our knowledge, this is the first study to demonstrate that nitrification, rather than denitrification, is the predominant microbial process of N2O production in situ in the study area in the tropical forest.