Genetic manipulation of baker's yeast for improved maltose utilisation

Two yeast/E.coli shuttle vector plasmids were studied in 1994, termed pIBIDB and pBP33. According to this study, each plasmid should contain at least one ADH2UAS (upstream activation sequence in the alcohol dehydrogenase 2 gene) insert. In the present study, the constructed plasmids were analysed and transformed into laboratory strain yeast. The aim of this project was to identify the orientation, quantity and quality of the insert in the selected plasmids. Methods such as restriction analysis, polymerase chained reaction (PCR), sequencing, plate assays and enzyme assays were used to identify and evaluate the novel inserts. The data presented in this thesis suggest the inserted ADH2UAS fragment did enhance the production of maltose permease and maltase when the transformants were cultivated in maltose and ethanol-glycerol medium. The results suggested that transformants containing two inserts of ADH2UAS had a greater influence on the transformants than a single insert. But the inserts within the vectors and in transformed laboratory stain yeast appeared unstable. This could be due to the method used for plasmid construction and the storage condition of the transformant