The ideal transgene vetors are expected to have no or low effect on normal cells but high transfection efficiency on carcinoma cells. In this study, the novel poly(diethylaminoethylmethacrylate)(PDEAEM)/PluronicŪ F127 pentablock copolymers were found to be able to mediate high-efficiency transfection of human epithelial ovarian carcinoma (SKOV3) cells while showing significantly lower efficiency of human epithelial retina (ARPE-19) cells an another animal normal cell line (swiss 3T3). To further examine the intracellular routes of polyplexes in SKOV3 and ARPE-19 cells, pentablock copolymers and plasmid DNA were covalently attached with Alexa FluorŪ 647 and Ethidium monoazide (EMA), respectively. Confocal microscopic images showed that plenty of polyplexes translocated into nuclei within 3h of transfection in SKOV3 cells, yet only few polyplexes were observed in APRE-cells. This difference in the amount of polyplexes in nuclei was also found in the cells at 10h and 24h posttransfection, indicating less nuclear entry may result in the lower efficiency of transfection. Since Brdu staining showed that SKOV3 proliferated much faster than ARPE-19 did, the nuclear entry of polyplexes was assumed to be correlated with the proliferation rate, and we hypothesized our pentablock copolymer could mediate gene delivery selectively in fast growing cells. Furthermore, in APRE-19 cells free DNA showed a weak signal or was localized around the cell membrane area, which implied that the uncomplexed DNA may have been degraded by the nuclease in the cytoplasm, or might have been exported out of the cell via exocytosis or other mechanisms. Although the validity of the hypothesis that our pentablock copolymer could selectively transfect hyperproliferative cells needs further examination, this present work provides a new perspective to design targeting vectors based on different characteristics among specific cells.