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Weeds compete with crops for growth resources, causing tremendous yield losses. Paraquat is one of the three most common non-selective herbicides. To study the mechanisms of paraquat resistance, we need to trace the movement of paraquat in plants and within the cell. 14C is a radioactive carbon isotope widely used to trace substances of interest in various biological studies, especially in transport analyses. Here, we describe a detailed protocol using 14C-paraquat to demonstrate paraquat efflux in Arabidopsis protoplasts.