An active targeting drug delivery system that targets the nucleus could solve the problem of the treatment of genetic disorders through gene delivery, but it has met with limited success. The purpose of this study was to establish an RNA aptamer-modified nucleus-targeting liposomal carrier system referred to as NupApt-liposomes. RNA aptamers against the Nup358 protein are prepared using a newly established Protein SELEX method. After confirming aptamer binding to the recombinant protein, an aptamer-lipid conjugate (Apt-PEG-DSPE) was prepared. Aptamer-modified liposomes and simple polyethylene glycol (PEG) liposomes were prepared to check its ability to bind to isolated nuclei. Confocal studies indicated that the aptamer-modified liposomes had the ability to bind to isolated nuclei, whereas PEG-liposomes showed only weak binding. Confocal laser scanning microscopy studies of inhibition assays also supported the above conclusion. The dissociation constant of the Nucleoporin358-specific aptamer referred to as NupApt01 and NupApt02 were 36 and 70 nM, respectively. Finally, with aptamer-modified liposomes, gene expression studies showed a two times better gene expression in NupApt-liposome-treated nuclei in comparison to that of PEG-liposomes. This represents the first artificial RNA aptamer-modified liposomes promoting the specific binding of a nanocarrier to the nucleus, thus improving gene expression in comparison to PEG-liposomes.