The use of non-viral vectors (plasmids) for therapeutic purposes is gaining increasing importance, as evidenced by the growing number of clinical evaluations using these vectors. Such vectors contain elements for replication in the host (bacteria) and elements for expressing the antigenic protein of interest. This allows us to estimate that in the near future the demand for plasmid DNA (pDNA) will substantially increase. pDNA production is carried out using Escherichia coli (E. coli) in high cell density cultures to maximize volumetric efficiency. In this review we analyze the recent work that has been done both in strains and in vectors to increase the production yield of plasmid DNA, in addition to facilitating purification.