Incorporating ligands with nanoparticle-based carriers for specific delivery of therapeutic nucleic acids (such as antisense oligonucleotides and siRNA) to tumor sites is a promising approach in anti-cancer strategies. However, nanoparticle-based carriers remain insufficient in terms of the selectivity and transfection efficiency. In this paper, we designed a dual receptor-targeted QDs gene carrier QD-(AS-ODN+GE11+c(RGDfK)) which could increase the cellular uptake efficiency and further enhance the transfection efficiency. Here, the targeting ligands used were peptides GE11 and c(RGDfK) which could recognize epidermal growth factor receptors (EGFR) and integrin ανβ3 receptors, respectively. Quantitative flow cytometry and ICP/MS showed that the synergistic effect between EGFR and integrin ανβ3 increased the cellular uptake of QDs carriers. The effects of inhibition agents showed the endocytosis pathway of QD-(AS-ODN+GE11+c(RGDfK)) probe was mainly clathrin-mediated. Western blot confirmed that QD-(AS-ODN+GE11+c(RGDfK)) could further enhance gene silencing efficiency compared to QD-(AS-ODN+GE11) and QD-(AS-ODN+c(RGDfK)), suggesting this dual receptor-targeted gene carrier achieved desired transfection efficiency. In this gene delivery system, QDs could not only be used as a gene vehicle but also as fluorescence probe, allowing for localization and tracking during the delivery process. This transport model is very well referenced for non-viral gene carriers to enhance the targeting ability and transfection efficiency.