Thesis (MSc. (Microbiology)) -- University of Limpopo, 2015 / S. cerevisiae has many properties which have made it the preferred host for the expression and production of a number of recombinant proteins. Xylose is the second most abundant sugar in nature and S. cerevisiae has been engineered to grow on this abundant sugar. Therefore, identifying S. cerevisiae promoters that are strongly induced during growth on xylose will be important in the production of recombinant proteins for the biofuel and other industries. Since xylose is not a native substrate for S. cerevisiae, it is not known how S. cerevisiae promoters will react during growth on xylose. The objective of the study was to evaluate the expression of a reporter gene, the Trichoderma reesei xylanase 2 (XYN2), under the control of five commonly used expression promoters (GPD3, ENO2, PGK1, ADH2 and YG100). Five episomal expression vectors were constructed for this purpose. These vectors were transformed to a recombinant xylose utilizing S. cerevisiae. Xylanase activity assays were used to determine the expression level from each of these promoters. The PGK1 promoter was observed to be the strongest promoter with average activity/OD of 130 nkat/ml/OD on both xylose and glucose. The GPD3 promoter showed the highest average activity/OD of 150 nkat/ml, but xylanase was only produced during growth on glucose. The data presented show that xylose is not a better carbon source than glucose for recombinant protein production in terms of the S. cerevisie promoters evaluated. Further research is required to obtain a yeast strain that grows well on xylose and promoters that show higher level on protein production.
Keywords: xylose, promoter, expression, recombinant, S. cerevisiae
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ul/oai:ulspace.ul.ac.za:10386/1758 |
| Date | January 2015 |
| Creators | Mande, Livhuwani |
| Contributors | La Grange, D. C., Van Zyl, W. H., Ncube, I. |
| Publisher | University of Limpopo |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | xi, 55 leaves |
| Relation |
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