Nonviral gene therapy has been hampered by its low transfection efficiency due to inefficient endocytosis, cysolic release, transnuclear location, and dissociation of DNA. Many polymeric carriers such as polyethyleneimine and poly-L-lysine rely on their proton buffering capacity called ‘proton sponge effect' which has been seriously questioned recently. Moreover, dissociation of condensed DNA from polymeric backbone, which is critical for cellular machineries to express newly introduced DNA, has not been fully understood. Significantly improved transfection of SW480 human colon cancer cells was achieved by delivering plasmid DNA in degradable nanoparticles which were synthesized by polymerizing cationic degradable monomers. Unlike other nonviral gene carriers using pre-synthesized polymers, the new approach provides control of molecular weight, composition, surface properties, and size of the carriers. The new carriers use swelling of the particles and osmotic destabilization to escape from the endosome rather than the questionable proton sponge effect. Moreover, cationic branches holding anionic DNA are cleaved from the polymeric backbone of the nanoparticles and make plasmid DNA accessible for efficient transcription and translation. Due to synthetic flexibility, functional moieties such as targeting ligands, monoclonal antibodies, and imaging tags, can be easily incorporated with the carriers. Design and synthesis as well as in vitro characterization of the nanoparticles, and enhanced ex vivo transfection confirmed by GFP and lacZ expression will be presented. In vivo transfection and imaging are also in progress.