Objective To synthesize a novel prostate cancer targeting gene vector PAMAM-PEG-C2min and improve gene transfection efficiency targeting on prostate cancer. Methods The aptamer (C2min) and polyamide-amine (PAMAM) were ligated by polyethylene glycol (PEG). The structure of the synthesized PAMAM-PEG-C2min was identified by NMR. The biological characteristics of the nanoparticles were examined by the uptake experiments and gene transfection experiments (the loaded gene was siR-M) with the prostate cancer cells (PC3 and LNCaP). Besides, the in vivo targeting was investigated using in vivo image system. The in vivo targeting results indicated that PAMAM-PEG-C2min can achieve the simultaneous targeting of two prostate cancer tissues. Results The PAMAM-PEG-C2min synthesis was confirmed by NMR. Cell uptake experiments showed that the cell uptake efficiency of PAMAM-PEG-C2min was concentration dependent. In vitro experiments showed that the PC3 and LNCaP cells transfection efficiency and targeting of PAMAM-PEG modified with C2min were significantly improved compared with the PEG modified PAMAM. Conclusion PAMAM-PEG-C2min is a potential targeted drug delivery vehicle. It provides a new technology platform for comprehensive and specific targeting treatment of prostate cancer.