Design, fabrication and testing of an electrochemically based DNA biosensor for detection of Coniella Granati fungal disease of pomegranate fruit and aster yellows bacterial grapevine disease

Madufor, Kennedy John Ndubuisi (2019-04)

Thesis (MEng)--Stellenbosch University, 2019.

Thesis

ENGLISH ABSTRACT: A portable low cost electrochemical DNA potentiostat biosensor was investigated for the detection of Coniella granati pathogenic disease of pomegranate fruit, using linear biotinylated DNA probes, immobilized on streptavidin modified screen printed carbon electrode. A cyclic voltammetric based electrochemical technique was chosen to detect the presence of Coniella granati pathogenic disease. A modified streptavidin screen printed carbon electrode was selected as a transducer, with the biotinylated probe DNA immobilized non-covalently on the electrode workspace. The streptavidin-biotinylated probe DNA was validated by the use of streptavidin coated magnetic beads, with the biotinylated probe DNA in a bind or wash buffer solution. A portable electrochemical potentiostat was designed and fabricated to electronically detect the change in current peak due to the electrochemical reduction and oxidation reaction of the DNA samples, after the immobilization occurred. A potassium ferricyanide solution was used to perform a functionality sensor test with the portable potentiostat. The expected change in current peaks due to the redox reaction of the sample was observed. The operation of the sensor and portable potentiostat was then verified by successfully measuring different DNA concentrations of aster yellows and Coniella granati. The results confirmed that the potentiostat biosensor is capable to detect plant disease successfully.

AFRIKAANSE OPSOMMING: 'n Draagbare, laekoste elektrochemiese DNS potensiostaat biosensor is ondersoek vir die opspoor van die siekte Coniella granati in pomogranate, deur gebruik te maak van lineêre DNS probes wat deur middel van biotin-streptavadin koppelings geïmmobiliseer word op gedrukte koolstof elektrodes. 'n Sikliese voltammetries-gebaseerde tegniek is gebruik vir die opspoor van die siekte Coniella granati. 'n Gedrukte koolstofelektrode, gemodifiseer met streptavadin, is as omsetter gekies, met DNS probes wat deur middel van biotin nie-kovalent gekoppel word aan die elektrode. Die streptavadin-biotin koppeling van die DNS probe is geverifieer deur gebruik te maak van magnetiese krale, wat bedek is met streptavadin, in 'n bufferoplossing. Die draagbare elektrochemiese potensiostaat is ontwerp en vervaardig om die verandering van piekstroom as gevolg van die reduksie-oksidasie reaksie waar te neem nadat DNS hibridisasie plaasgevind het. 'n Kalium ferri-sianied oplossing is vervolgens gebruik om die funksionaliteit van die sensor met die draagbare potensiostaat te toets. Die verwagte vernadering in stroompieke as gevolg van die redoks reaksies van die steekproewe is waargeneem. Die werking van die sensor en die draagbare potensiostaat is geverifieer deur verskillende DNS konsentrasies van aster yellows en Coniella granati suksesvol te meet. Die resultate bevestig dat die potensiostaat biosensor in staat is om deteksie van plantsiektes suksesvol te doen.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/105885
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