Nanoparticles formulated from polylactic-co-glycolic acid (PLGA) polymer loading a new recombinant plasmid pEGFP-TKAFB (TK-PLGA-NPs) were prepared by a double-emulsion evaporation technique. Both in vitro and in vivo release behaviors of TK-PLGA-NPs (with particle diameter ranged from 50 to 100 nm) were investigated, using ethidium bromide (EB) staining and gamma scintigraphy, respectively. The results indicated that the in vitro release rate of DNA (pEGFP-TKAFB plasmid) in TK-PLGA-NPs showed good fit into the Higuichi Equation and dependence in the molecular weight of PLGA polymer. 0.5 h after injection of nanoparticles containing (32)P labeled pEGFP-TKAFB plasmid ((32)P-TK-PLGA-NP) via caudal vein of the mice, the ratio of radioactivity intensity in the liver to total intensity was above 70%, which showed a 1.4-fold increase over that by injection of (32)P labeled pEGFP-TKAFB plasmid ((32)pEGFP-TKAFB plasmid, (32)P-TK). Similarly, 2 h after hypodermic injection of (32)P-TK-PLGA-NPs in mice, the ratio of radioactivity in the liver against total radioactivity was more than 70%, which was 1.6-fold compared with naked (32)P-TK. All these data showed that the TK-PLGA-NPs has the potential for liver-targeting and delayed drug release.