The manufacturing of plasmid vectors for gene therapy applications depends on the yield of plasmid DNA production, which has to be high enough to provide sufficient amount of super coiled plasmid DNA. In the downstream process the amount and quality of plasmid DNA are affected by many factors, which have to be optimised in the upstream process of bioprocess development. The scope of the thesis was to increase the yield of the upstream process. Bacterial cultivation began in a 3 L bioreactor vessel where the growth of bacteria, pH and oxygen partial pressure were monitored in a controlled environment. Bacterial cultivation was carried out a batch mode and also in a fed-batch mode. The effect of oxygen partial pressure and temperature on the production of plasmid DNA was tested. The use of fed-batch mode increased the yield of plasmid DNA by 79.5% compared to batch mode.