Abstract: Clarifying the effects of Chinese milk vetch (CMV) incorporation and substitution of conventional nitrogen fertilizers with slow-release nitrogen fertilizers on the soil organic carbon (SOC) fractions, carbon pool management index (CPMI), and maintenance/increase in rice yield can provide a theoretical basis for formulating application schemes for slow-release nitrogen fertilizers that are conducive to SOC sequestration and sustainable utilization of paddy fields. In this study, the effects of CMV incorporation and substitution of conventional nitrogen fertilizers with slow-release nitrogen fertilizers on the crop yield, economic benefit, SOC and labile organic carbon (LOC) content, and CPMI were studied quantitatively in rice-wheat and rice-CMV rotation systems in the middle and lower reaches of the Yangtze River, and their effects on the maintenance/increase in rice yield were investigated. The experiment adopted a two-factor, completely randomized block design, with the primary factors being the crop rotation systems rice-wheat (W-R) and rice-CMV (CMV-R). The secondary factors were nitrogen fertilizer application methods, including conventional nitrogen fertilizer application (CN), slow-release nitrogen fertilizer substitution for conventional nitrogen fertilizer based on the same nitrogen input (SRN), 20% nitrogen reduction during the rice season and 10% nitrogen reduction during the wheat season based on SRN (T1), 40% nitrogen reduction during the rice season and 20% nitrogen reduction during the wheat season based on SRN (T2), and 60% nitrogen reduction during the rice season and 30% nitrogen reduction during the wheat season based on SRN (T3). Compared with the W-R rotation system, rice yield of the CMV-R rotation system significantly increased by 2.1% (P<0.05), whereas the annual net income significantly decreased by 20.4% (P<0.05). Compared with the W-R rotation system, the total LOC (TLOC), high LOC (HLOC), and medium LOC (MLOC) content as well as the CPMI of the CMV-R rotation system significantly increased by 5.2%, 6.0%, 5.1%, and 6.6% (P<0.05), respectively. Compared with the CN treatment, the rice yields under the SRN and T1 treatments significantly increased by 12.6% and 7.3% (P<0.05), respectively, and annual net incomes significantly increased by 38.8% and 20.7% (P<0.05), respectively, whereas the rice yield under the T3 treatment significantly decreased by 9.4% (P<0.05). Compared with the CN treatment, the TLOC and HLOC content, and CPMI under the SRN treatment significantly increased by 7.1%, 6.9%, and 9.4% (P<0.05), respectively. Based on the crop rotation system and nitrogen fertilizer application method, the rice yield, SOC content, TLOC content, and CPMI under the SRN treatment and the CMV-R rotation system were the highest, followed by the SRN treatment under the W-R rotation system and the T1 treatment under the CMV-R rotation system. No significant difference was found for these indicators between the T1 treatment under the CMV-R rotation system and the SRN treatment under the W-R rotation system. Therefore, in rice-growing regions in the middle and lower reaches of the Yangtze River, adopting a rice-CMV rotation system combined with slow-release nitrogen fertilizers to replace conventional nitrogen fertilizer (SRN), and even replacing it with a 20% reduction in the rice season based on SRN, is conducive to SOC sequestration and the sustainable utilization of paddy fields, which is of great significance for sustainable agricultural production in this region.