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Optimization of purification and characterisation of over-expressed rotavirus capsid protein VP6

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dc.contributor.advisor Gildenhuys, Samantha
dc.contributor.advisor Parbhoo, Nishal
dc.contributor.author Kgokolo, Samuel Maphalle
dc.date.accessioned 2018-04-10T06:38:41Z
dc.date.available 2018-04-10T06:38:41Z
dc.date.issued 2017-12
dc.identifier.citation Kgokolo, Samuel Maphalle (2017) Optimization of purification and characterisation of over-expressed rotavirus capsid protein VP6, University of South Africa, Pretoria, <http://hdl.handle.net/10500/23726>
dc.identifier.uri http://hdl.handle.net/10500/23726
dc.description.abstract Rotavirus is responsible for the death of many children annually, and current vaccines have lower efficiency in developing countries. A reverse translated consensus gene sequence of the rotavirus VP6 cloned into a pET-28a(+) plasmid was used to transform BL21 and KRX Escherichia coli cells. Optimal expression of soluble protein was induced in KRX cells by adding 0.05% L-rhamnose and 0.0001 M IPTG, with an incubation temperature of 25ºC for 6 h. VP6 was purified by combining anion exchange chromatography followed by affinity chromatography. Far-UV circular dichroism and intrinsic fluorescence were used as probes to assess the native structure of VP6 and structural in the presence of a denaturant, high sodium chloride concentrations and varying temperatures. The 0.2 M sodium chloride had an impact on the VP6’s tertiary structure and also influenced the proteins conformational changes as detected during thermal unfolding to 90ºC. Although treatment with 3 M urea showed tertiary structural changes no secondary structural loss occurred due to the presence of a denaturant. en
dc.format.extent 1 online resource (xii, 99 leaves) ; illustrations (some color) en
dc.language.iso en en
dc.subject Chromatography en
dc.subject Circular dichroism en
dc.subject Escherichia coli en
dc.subject Fluorescence en
dc.subject Plasmid en
dc.subject Protein conformation en
dc.subject Protein expression en
dc.subject Purification en
dc.subject Rotavirus en
dc.subject VP6 en
dc.subject.ddc 616.926
dc.subject.lcsh Rotavirus infections en
dc.subject.lcsh Molecular epidemiology en
dc.subject.lcsh Chromatographic analysis en
dc.subject.lcsh Gastritis -- Microbiology en
dc.subject.lcsh Escherichia coli infections en
dc.title Optimization of purification and characterisation of over-expressed rotavirus capsid protein VP6 en
dc.type Dissertation en
dc.description.department Life Sciences en
dc.description.degree M. Sc. (Life Sciences) en


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  • Unisa ETD [12519]
    Electronic versions of theses and dissertations submitted to Unisa since 2003

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