Einsatz von Pflanzenzellkulturen als industriell nutzbare Expressionssysteme für therapeutische Proteine

Within the present thesis transgenic BY-2 cultures should be tested regarding their potential as industrial expression system for therapeutic proteins. For this purpose, BY-2 cultures expressing recombinant HSA under control of the CaMV 35SS promoter were developed. A leader peptide ensured the secretion of the protein to the apoplast. Main focus of the described studies are such parameters, which are of essential importance for the industry, like productivity, genetic stability of the organism as well as the production of a functional protein and the economy of the production process. In general the reproducibility of developed and optimised industrial production processes are ensured by cell stocks of the relevant organisms. Such cell banks guarantee especially the genetic stability of initially used organisms. Traditionally periodic subculturing in shake flasks performs the storing of plant cell cultures. Thus the cells are weekly supplied with fresh medium to maintain their vitality. During this kind of maintenance changes regarding expression and productivity of recombinant proteins could be detected. Hence the development of alternative storing opportunities is of crucial relevance for the industry. Within these studies a protocol for the long-time storage of transgenic BY-2 cultures was developed. In the process development media optimisation is a helpful tool to increase the productivity of the used organisms. Especially for plant cell cultures commercialised media are overaged and not defined for single cell lines. This indicates the enormous room of improvement, which could also be shown within these studies: by variation of the concentration of only one media component the extracellular HSA amount could be increased 17-fold. All results generated during shake flasks trials could be transferred to a larger scale performed in stirred tank bioreactors. The combination of two different enhancing substances did not yield in a summation of the effects during shake flask trials. In stirred tank bioreactors the combination of two enhancing substances increased the productivity by a factor of 20. Even if these results correspond to data, which were generated in other studies performed at the IME (data not shown), a generalization about the production of recombinant proteins in plant cell cultures is not allowed yet. The development and optimisation of a fermentation strategy for the effective production of recombinant proteins in plant cell cultures was an important part of this thesis. Different fermentation strategies, their variations as well as different process parameters should be investigated. All process strategies of industrial interest like the conventional batch fermentation as well as the continuous fermentation were applicable. An outstanding result was the continuous fermentation: during cultivation time of 50 days an extensively constant amount of HSA could be daily harvested. This thesis was supported by the RWTH Aachen Postgraduate Sponsorship for Young Scientists.

Grantor dc:publisher

Publikationsserver der RWTH Aachen University

Year dc:date

2007

Author dc:creator

• Schmale, Kathrin

Contributors dc:contributor

• Fischer, Rainer

• info:eu-repo/classification/ddc/570
• Kontinuierliche Fermentation
• Kryokonservierung
• Humanes Serumalbumin
• Biowissenschaften, Biologie
• BY-2 Kulturen
• Medienoptimierung
• rekombinante Proteine
• Pflanzenzellkulturen
• Expressionssystem
• By-2 cultures
• media optimisation
• recombinant proteins
• plant cell cultures
• expression systems

Statement dc:rights

• info:eu-repo/semantics/openAccess

Language dc:language

ger

Identifier

https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123981%22

OAI identifier oai:identifier

oai:publications.rwth-aachen.de:62410