Based on a long-term fertilization experiment in a paddy soil, this paper studied the effects of long-term application of chemical fertilizer and rice straw on carbon (C) and nitrogen (N) mineralization by incubation experiments. Results showed that long-term application of chemical fertilizer and straw increased carbon mineralization, but decreased the proportion of C mineralized to total organic C. Long-term application of chemical fertilizer increased N mineralization and the proportion of mineralizable N to total N. Compared to chemical fertilizer application alone, added application of rice straw did not increased N mineralization. Long-term application of chemical fertilizer and rice straw significantly increased microbial biomass C and N contents.