As fossil fuels are depleted worldwide, alternative resources is required to replace fossil fuels, and biofuels are in the spotlight as alternative resources. Biofuels are produced from biomass, which is a renewable resource to produce biofuels or bio-chemicals. Especially, in order to substitute fossil fuels, the research focusing the biofuel (biodiesel) production based on CO2 and biomass achieves more attention recently. To produce biomass-based biodiesel, the development of a supply chain network is required considering the amounts of feedstocks (ex, CO2 and water) required producing biodiesel, potential locations and capacities of bio-refineries, and transportations of biodiesel produced at biorefineries to demand cities. Although many studies of the biomass-based biodiesel supply chain network are performed, there are few types of research handled the uncertainty in CO2 supply which influences the optimal strategies of microalgae-based biodiesel production. Because CO2, which is used in the production of microalgae-based biodiesel as one of important resources, is captured from the off-gases emitted in power plants, the uncertainty in CO2 supply from power plants has big impacts on the optimal configuration of the biodiesel supply chain network. Therefore, in this study, to handle those issues, we develop the two-stage stochastic model to determine the optimal strategies of the biodiesel supply chain network considering the uncertainty in CO2 supply. The goal of the proposed model is to minimize the expected total cost of the biodiesel supply chain network considering the uncertain CO2 supply as well as satisfy diesel demands at each city. This model conducted a case study satisfying 10% diesel demand in the Republic of Korea. The overall cost of the stochastic model (US$ 12.9/gallon·y) is slightly higher (23%) than that of the deterministic model (US$ 10.5/gallon·y). Fluctuations in CO2 supply (stochastic model) had a significant impact on the optimal strategies of the biodiesel supply network.