Plasmid DNA fermentation strategies: influence on plasmid stability and cell physiology |
| |
Authors: | Filomena?Silva Jo?o?A?Queiroz Email author" target="_blank">Fernanda?C?DominguesEmail author |
| |
Institution: | 1.CICS-UBI, Health Sciences Research Centre,University of Beira Interior,Covilh?,Portugal;2.Centro de Investiga??o em Ciências da Saúde,Universidade da Beira Interior,Covilh?,Portugal |
| |
Abstract: | In order to provide sufficient pharmaceutical-grade plasmid DNA material, it is essential to gain a comprehensive knowledge
of the bioprocesses involved; so, the development of protocols and techniques that allow a fast monitoring of process performance
is a valuable tool for bioprocess design. Regarding plasmid DNA production, the metabolic stress of the host strain as well
as plasmid stability have been identified as two of the key parameters that greatly influence plasmid DNA yields. The present
work describes the impact of batch and fed-batch fermentations using different C/N ratios and different feeding profiles on
cell physiology and plasmid stability, investigating the potential of these two monitoring techniques as valuable tools for
bioprocess development and design. The results obtained in batch fermentations showed that plasmid copy number values suffered
a pronounced increase at the end of almost all fermentation conditions tested. Regarding fed-batch fermentations, the strategies
with exponential feeding profiles, in contrast with those with constant feeding, showed higher biomass and plasmid yields,
the maximum values obtained for these two parameters being 95.64 OD600 and 344.3 mg plasmid DNA (pDNA)/L, respectively, when using an exponential feed rate of 0.2 h−1. Despite the results obtained, cell physiology and plasmid stability monitoring revealed that, although higher pDNA overall
yields were obtained, this fermentation exhibited lower plasmid stability and percentage of viable cells. In conclusion, this
study allowed clarifying the bioprocess performance based on cell physiology and plasmid stability assessment, allowing improvement
of the overall process and not only plasmid DNA yield and cell growth. |
| |
Keywords: | |
本文献已被 PubMed SpringerLink 等数据库收录! |
|