The biochemical properties of soil have often been described as early and sensitive indicators of ecological changes in both natural soil and agroecosystem. In the current study, the impacts of the amendment of Bt-transgenic rice (KMD) straw on biological activities in water-flooded soil were investigated under laboratory conditions and compared with non-transgenic rice (Xiushui 11) straw. The results showed that there were some differences in protease, neutral phosphatase and cellulase activities between soil amended with Bt-transgenic rice straw and non-transgenic rice straw at the early stage of incubation, and none of these differences were persistent. However, differences in dehydrogenase activity, methanogenesis, hydrogen production and anaerobic respiration between soil supplemented with Bt-transgenic rice straw and non-transgenic rice straw were persistent over the course of incubation. Dehydrogenase activity, methanogenesis and anaerobic respiration were considerably lower from sample days 7to 56, but higher after day 56in soil amended with Bt-transgenic rice straw. In comparison, the H2-production in soil containing Bt-transgenic rice straw was significantly lower after day 56. The results demonstrated that the amendment of the Bt-transgenic rice straw altered some important biological properties in water-flooded soil, indicating a shift in microbial populations or a change in the metabolic abilities of the microbial community as a result of substrate availability in soil.