The purpose of this paper was to establish the surface modified poly(d,l-lactide-co-glycolide) (PLGA) nanosphere platform with chitosan (CS) for gene delivery by using the emulsion solvent diffusion (ESD) method. The advantages of this method are a simple process under mild conditions without sonication. This method requires essentially dissolving both polymer and drug in the organic solvent. Therefore a hydrophilic drug such as nucleic acid is hardly applied to the ESD method. Nucleic acid can easily form an ion-complex with cationic compound, which can be dissolved in the organic solvent. Thereafter, nucleic acid solubility for organic solution can increase by complexation with cationic compound. We used DOTAP as a cationic compound to increase the loading efficiency of nucleic acid. By coating the PLGA nanospheres with CS, the loading efficiency of nucleic acid in the modified nanospheres increased significantly. The release profile of nucleic acid from PLGA nanospheres exhibited sustained release after initial burst. The PLGA nanospheres coated with chitosan reduced the initial burst of nucleic acid release and prolonged the drugs releasing at later stage. Chitosan coated PLGA nanosphere platform was established to encapsulate satisfactorily wide variety of nucleic acid for an acceptable gene delivery system.