DNA vaccines and gene medicines, derived from bacterial plasmids, are emerging as an important new class of pharmaceuticals. DNA molecules made strictly by rational design allows the manufacturer to bypass years of development for the production of efficacious vaccines, and literally create new vaccine entities in days and mass produce vaccines in 2-3 weeks for rapid deployment against new biological agents. However, existing fermentation and purification production technologies are inefficient, difficult to scale, and require sophisticated production equipment and methodology. This limits their utility to meet cost and capacity needs for existing plasmid applications, or to rapidly produce kilograms of plasmid DNA for pandemic vaccination. Phase I addressed these problems directly and successfully by proving feasibility of a cost effective dramatically streamlined and simple purification process that eliminates specialized and costly alkaline or heat lysis steps and the associated toxic waste streams. In Phase II, we will make and evaluate designer strains and associated fermentation and purification methodology. These strains will utilize novel chimeric nucleases and autolytic bacterial strains developed in Phase I. Four prototypes are envisioned. Prototype I is a rapid deployment plasmid production system linking autolytic plasmid purification to Nature Technology Corporation's (NTC) existing fermentation platform to facilitate immediate production of a variety of plasmid DNAs for pandemic applications. The key drivers for this platform are speed of production, safety, simplicity, scalability and transferability to existing manufacturing facilities in developed and emerging countries. Autolytic strains with an integrated chimeric nuclease will be created, and fermentation, autolysis, and plasmid purification methodologies developed. The system will be integrated into NTC's existing rapid deployment DNA vaccine platform (RapidVACC) for response to influenza H5N1 pandemic. Prototypes II-IV are aimed at facilitating the commercialization of gene medicines for a variety of applications. The key drivers for these prototypes are purity, yield, cost (reagents, equipment, volumes, and time), scalability and safety. Prototype IV links high yield defined media fermentation with new simplified lysis and purification methodologies. This technology will be licensed to plasmid DNA manufacturers; one such commercialization channel has already been identified. [unreadable] [unreadable] [unreadable]