Brassica napus is a Cd hyperaccumulator, which is a serious threat to food and fodder safety. However, no related studies on developing Cd-safe B. napus have been reported yet. Here, we screened out a novel Cd uptake-related gene, AtCUP1, from the major facilitator superfamily in Arabidopsis thaliana . The mutation of AtCUP1 decreased Cd accumulation, both in roots and shoots of A. thaliana . Furthermore, the disruption of the AtCUP1 gene by the CRISPR/Cas9 system significantly reduced Cd accumulation in A. thaliana . Interestingly, the disruption of the BnCUP1 gene, an orthologous gene of AtCUP1 , by the CRISPR/Cas9 system also diminished Cd accumulation in both roots and shoots of B. napus based on the hydroponics assay. Furthermore, for the field experiment, the Cd accumulations of BnCUP1 -edited lines were reduced by 52% in roots and 77% in shoots compared to that of wild-type (WT) lines, and the biomass and yield of BnCUP1 -edited lines increased by 42% and 47% of that of WT, respectively. Noteworthily, agronomic characteristics of B. napus were not apparently affected by BnCUP1 -editing. Thus, BnCUP1 -edited lines are excellent non-transgenic germplasm resources for reducing Cd accumulation without a distinct compromise in yield, which could be applied to agricultural production in Cd-contaminated soils.