Abstract: A agricultural ecosystem is a significant source of greenhouse gas emission, and the development of low-carbon agriculture represents a promising approach to the mitigating of climate change. Given the substantial economic disparities between the northern and southern regions of Jiangsu Province, implementing region-specific carbon reduction measures is essential to maximizing emission reduction in agriculture and promoting sustainable development. This study, based on survey data from farmers, focuses on the carbon footprint of the rice-wheat annual rotation system in Jiangsu Province, investigating the regional differences between the northern and southern parts of the province. A life cycle assessment (LCA) methodology was employed to systematically collect and analyze agricultural production and carbon emission data from Huaian and Suzhou cities in Jiangsu. The results reveal significant differences in the carbon footprints of rice and wheat production between Suzhou and Huaian (P < 0.05). In rice cultivation, methane (CH₄) emissions were the primary source of the carbon footprint, contributing 67.3%. In contrast, nitrogen fertilizer usage accounted for the largest share of the carbon footprint in wheat cultivation (55.5%), followed by nitrous oxide (N₂O) emissions (25.3%). Additionally, the scale of rice and wheat cultivation have significant impact on the carbon footprint. Both rice and wheat production processes demonstrate that nitrogen fertilizer application and diesel consumption were the main factors influencing the carbon footprint, with both showing a positive correlation with emissions. There were notable regional differences in the carbon footprint of the rice-wheat annual rotation system between the northern and southern regions of Jiangsu Province, with carbon emissions in Suzhou being significantly lower than in Huaian. Therefore, when conducting carbon footprint assessments of agricultural production systems in Jiangsu, it is important to consider the close relationship between economic development and agricultural production models. Such an approach will contribute to the formulation of region-specific policies for sustainable agricultural development and provide a foundation for the establishment of a low-carbon agricultural production system.