Vaccines have proven to be an effective means to protect communities from a range of human and agricultural pathogens. Unfortunately, costs associated with the development and manufacturing of vaccines often prevent some of the neediest populations from receiving and distributing these essential prophylactics. Advances in molecular and synthetic biology represent potential low cost solutions for enhancing bioproduction. In the following chapter, we describe a program in which RNA Interference (RNAi) has been successfully employed to identify gene modulation events that enhance poliovirus production in vaccine manufacturing cell lines. Transition of this technology into stable production lines promises to increase overall vaccine manufacturing capabilities – thereby making these essential, life-saving therapeutics available at an affordable cost.