Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.791136
Title: Metabolic flux balance application to plant cell cultures
Author: Yu, Hao
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2017
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Abstract:
In this research, a computational metabolic flux balance analysis was developed in order to investigate the biosynthetic potential of plant cell suspension cultures to produce heterologous proteins. It involved both theoretical/computational and experimental work. The model plants were Nicotiana tabacum, tobacco, the wild type and two genetically modified cell lines of tobacco carrying the genes coding for either β-Glucuronidase (GUS) or human transforming growth factor-β3 (TGFβ3) in their chloroplast genomes. The wild type and genetically modified tobacco seeds were germinated in order to initiate first callus formation and then the cell suspension cultures. These were used in batch experiments using Murashige-Skoog medium, in order to monitor growth, sugar consumption and heterologous protein production, for the first time according to the literature. The typical batch lasted 10-15 days, the cell concentration reached approximately 12-14 g dry weight /l for the wild type and 8-10 g dry weight /l for the genetically modified cell lines. The maximum specific growth rates were 0.0067 h-1 and 0.009 h-1, for the wild type and genetically modified cell lines, respectively. Experimental data were used in order to deveop kinetic models of growth and sugar uptake for both wild type tobacco and genetically modified tobacco cell cultures. The computational metabolic model involved 182 reactions and 149 metabolites which was then constructed in matrix formalism for the optimisation of a chosen objective function such as, the specific growth or heterologous protein production rate, in The General Algebraic Modelling System (GAMS) platform. Again, this was the first time such models were developed for such heterologous protein production systems. Although the experimental concentrations of these proteins were very low in the cell samples, the colour tints in the media indicated that they might have been secreted in to the medium which was not analysed. These experiments were not optimised in terms of medium composition and growth conditions for the maximisation of the product. The metabolic model indicated that plant cell cultures had the potential to produce as much as 55 g TGFβ3 protein/l and 289 g GUS protein/l.
Supervisor: Pandiella, Severino ; Mavituna, Ferda Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.791136  DOI: Not available
Keywords: kinetic model ; TGFß3 ; GUS ; plant cell ; metabolic flux balance ; tobacco
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