Straw incorporation generally increases CH<sub>4</sub> emission from rice fields, but its effects on the mechanism of CH<sub>4</sub> emission, especially on the pathway of CH<sub>4</sub> production and the fraction of CH<sub>4</sub> oxidized are not well known. To investigate the methanogenic pathway, the fraction of CH<sub>4</sub> oxidized as well as the stable carbon isotope fractionation during the oxidation and transport of CH<sub>4</sub> as affected by straw incorporation, production and oxidation of CH<sub>4</sub> in paddy soil and rice roots and δ<sup>13</sup>C-values of produced CH<sub>4</sub> and CO<sub>2</sub>, and emitted CH<sub>4</sub> were observed in incubation and field experiments. Straw incorporation significantly enhanced CH<sub>4</sub> production potentials of the paddy soil and rice roots. However, it increased the relative contribution of acetate to total CH<sub>4</sub> production (F<sub>ac</sub>) in the paddy soil by ~ 10–30%, but decreased F<sub>ac</sub>-value of the rice roots by ~ 5–20%. Compared with rice roots, paddy soil was more important in acetoclastic methanogenesis, with F<sub>ac</sub>-value being 6–30% higher. Straw incorporation highly decreased the fraction of CH<sub>4</sub> oxidized (F<sub>ox</sub>) by 41–71%, probably attributed to the fact that it increased CH<sub>4</sub> oxidation potential whereas CH<sub>4</sub> production potential was increased to a larger extent. There was little CH<sub>4</sub> formed during aerobic incubation, and the produced CH<sub>4</sub> was more <sup>13</sup>C-enriched relative to that of anaerobic incubation. Assuming δ<sup>13</sup>C-values of CH<sub>4</sub> aerobically produced in paddy soil to be the δ<sup>13</sup>C-values of residual CH<sub>4</sub> after being oxidized, F<sub>ox</sub>-value still appeared to be 45–68% lower when straw was incorporated. Oxidation fractionation factor (α<sub>ox</sub>) was higher with straw incorporation (1.033) than without straw incorporation (1.025). The δ<sup>13</sup>C-values of CH<sub>4</sub> emitted after cutting of the plants (−50–−43‰) were more positive than those of before (−58–−55‰), suggesting a transport fractionation factor (ϵ<sub>transport</sub>) was −8.0‰ with straw incorporation and −12.0‰ without straw incorporation. Reasons for this difference may be related to the decrease in growth of the rice crop as a result of straw incorporation. The experiment shows that straw incorporation increases the contribution of acetate to total methanogenesis in paddy soil but decreases it on rice roots, and it significantly decreases the fraction of CH<sub>4</sub> oxidized in the field, and expands oxidation fractionation while reducing transport fractionation.