ABSTRACT
Recent advances in gene therapy and nucleic acid vaccinations have resulted in an increased demand for plasmid DNA (pDNA). In this chapter, the authors discuss pDNA production by looking at the upstream process, the downstream process, and analytical development. The upstream process discusses plasmid optimization, media composition, and fermentation conditions that could be used during the production of pDNA. The downstream process explores cell disintegration approaches that can be applied, RNA and Endotoxin removal, clarification of lysate, chromatographic purification, formulation, and final storage of the product.
The analytical development section discusses methods that can address the regulatory requirements of pDNA product such as quantity, purity, identity, potency, quality, and safety. The methods that are used for the analysis such as A260, A260/A280 ratio, agarose gel electrophoresis (AGE), high pressure liquid chromatography (HPLC), and capillary electrophoresis utilizing a laser induced fluorescent detector (CE-LIF) have been discussed. In this chapter, a summary of the current knowledge of producing high yield pDNA that meets the quality, safety, and efficacy standards are discussed.
