Manure‐based biochar is an effective amendment to increase both carbon sequestration and the fertility of degraded soils, whereas the responses of N2O emissions and microbial‐N cycling genes to its application and the underlying mechanisms are poorly understood. Here, four biochars were produced under two pyrolysis temperatures (300 and 700°C) and with two organic C extraction procedures (water and acetone extraction). The resulting biochars were either with relative enrichment or depletion in easily mineralizable carbon (EMC) compared with recalcitrant C. We added biochars, with urea and sodium nitrate, to a degraded red soil and incubated the amended soils at moisture levels of 60% and 130% field capacity. All the biochars decreased nitrification gene abundance, that is, amoA. Biochars with EMC had greater stimulatory effects on the abundance of denitrification genes (nirK, nirS, and nosZ) and N2O emission, regardless of moisture level and N form, compared with biochar without EMC. Biochar‐induced microbial activity, biochar aliphatics, and alkyl groups correlated positively with N2O emission and denitrification gene abundance. The water dissolved organic C of biochar facilitated the conversion of N2O to N2, whereas the acetone extractable C fraction postponed the completion of denitrification. In conclusion, the N2O emission and denitrification gene abundance increased with decreasing biochar aromaticity and increasing EMC content. Our study emphasizes that the EMC supply in manure‐based biochars also importantly mediates soil N2O emission and microbial N cycling genes, adding to current understanding of influencing factors such as biochar pH, porosity, N availability, and redox property.