Poly(lactic-co-glycolic acid) (PLGA) particles have been widely explored as vehicles for delivery of plasmid DNA to mammalian cells both in vitro and in vivo. Achieving high incorporation efficiencies and control over release kinetics are significant challenges in encapsulating hydrophilic molecules such as DNA within submicron particles fabricated from PLGA. This study explored two modifications in the preparation of submicron particles to specifically address these challenges. Firstly, we compared homogenization and sonication as energy sources for emulsification. It was demonstrated that particles prepared with homogenization resulted in higher encapsulation efficiency and a linear release profile of DNA as compared to particles prepared with sonication, which exhibited lower encapsulation efficiency and a burst release. Also investigated was conjugation of poly-L-lysine to PLGA (PLGA-PLL) to create an electrostatically favorable interaction between the carrier material and the DNA. Particles fabricated with high weight percentages of PLGA-PLL/PLGA resulted in remarkably increased loading (>90%). Additionally, the release profile could be dictated by the quantity of PLGA-PLL incorporated into the particles. Particles incubated in vitro on COS-7 cells were able to transfect cells. These results demonstrated that DNA encapsulation and release were modulated by the method of fabrication.