Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.487493
Title: Green fluorescent protein as an analytical tool to dissect the physiology of recombinant protein production in fermenters
Author: Jones, Joanna Jane
Awarding Body: University of Birmingham
Current Institution: University of Birmingham
Date of Award: 2007
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Abstract:
A sensor exploiting the properties of Green Fluorescent Protein (GFP) was developed and was used to optimise fennentation conditions for production of the soluble fusion protein CheYGFP. A 9-fold increase in yield was achieved. Genes encoding CheY, Defl and DXR were fused in-frame to the 5' and 3' end ofthe GFP gene to detennine whether this affected optimised production of soluble GFP fusion proteins. Different conditions were required for optimal accumulation of the recombinant protein and the optimised conditions for CheYGFP and DXRGFP were identical to those for CheY and DXR respectively. Conditions optimal for the accumulation of Defl, DeflGFP, GFPDefl, GFPCheY, GFPDXR and GFP were the same. During several fennentations the production of soluble recombinant protein did not increase after 12 h. This was concomitant with a rapid increase in the rate of carbon dio~ide production by the culture. Cell pellets after centrifugation contained a layer of bacteria that were not fluorescent, indicating the presence of a non-productive population of bacteria. Restriction mapping and sequence analysis confinned the presence after 12 h of bacteria carrying a plasmid in which a frameshift mutation had occurred. Improvement in the maintenance of the original plasmid was achieved by further manipulation offennentation conditions.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.487493  DOI: Not available
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