Abstract: Exploring the mechanisms underlying high-yield formation and carbon balance in rice ratooning is important for further enhancing yield potential. This study was conducted in Putian, Fujian Province (25°18′N, 119°7′E). Two varieties with varying growth durations were used to examine the yield formation of the main crop (MC), ratooning season rice (RSR), and corresponding single-cropping rice (LR). The dynamics of rhizosphere microorganisms and net ecosystem carbon balance (NECB) in paddy fields, as well as the underlying mechanisms influencing these processes, were investigated in 2023 and 2024. The average daily yield for one season of RSR was 46.37%–82.66% and 42.35%–86.61% higher than that of MC and LR, respectively. This discrepancy was attributed to the 41.49% and 41.44% higher dry matter translocation efficiency of stems and leaves of RSR, and the greater amount of dry matter transferred to the panicles compared with MC and LR, respectively. Compared with those of MC and LR, the allocation amount of photosynthetic products to the below-ground parts (roots and soil) of RSR decreased by 56.05%–58.79% and 52.14%–53.19%, respectively, while the corresponding allocation proportions were reduced by 49.39%–54.11% and 49.28%–49.40%, respectively. Conversely, compared with those in MC and LR, the allocation of photosynthetic products to the panicles of RSR increased by 5.10%–17.67% and 19.48%–20.30%, respectively, with the corresponding allocation proportions increased by 21.04%–31.04% and 27.49%–29.14%, respectively. This, in turn, resulted in a higher harvest index and average daily yield in RSR. Moreover, analysis of greenhouse gases emission intensity and NECB in paddy fields demonstrated that the average daily greenhouse gases emission intensity under ratoon rice cultivation pattern decreased by 52.12%–60.96% compared with that under single-cropping rice cultivation pattern. Comparative analysis revealed significant differences in carbon sink capacities between cultivation patterns. The ratoon rice cultivation pattern exhibited robust carbon sequestration performance, achieving a carbon balance of 17.78–32.85 t(CO₂-eq)·hm⁻². In contrast, the single-cropping rice cultivation pattern showed relatively weaker carbon sink capacity with a surplus of only 5.70–11.67 t(CO₂-eq)·hm⁻². This divergence arises from two key mechanisms: 1) effectively extending the utilization period of light and thermal resources through a unique MC–RSR cycle configuration, and 2) modifying the soil microbial community by increasing carbon-fixing microbes while reducing carbon decomposers in the rhizosphere soil of RSR compared to MC and LR. These findings provide evidence to advance agricultural carbon neutrality technologies and achieve the goals of low-carbon, green, and sustainable agricultural development.