Theses and Dissertations (School of Molecular and Cell Biology)

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De novo production of Taxol intermediates by Saccharomyces Cerevisiae
(2022) Rabie, Renée
Taxol is an invaluable anticancer molecule produced by Yew trees and their endophytic fungi. Harvesting taxol is difficult and often has low yields. For these reasons, a method to produce taxol heterologously in a fast-growing, well-studied, safe microbe is desirable. In this study, artificial genes were designed for the expression of two taxol biosynthesis pathway enzymes, as well as an assisting NADPH-cytochrome P450 reductase. The genes were designed to be compatible with the pCut transformation technique, which allows genomic integration into Saccharomyces cerevisiae strain BY4742. The genes were then divided into seven fragments. Two additional DNA fragments were amplified directly from the yeast genome because their complexity made them difficult and expensive to synthesise. These nine DNA fragments were designed to be assembled into three linear fragments of equal length for transformation of S. cerevisiae. Attempts at assembling these nine fragments into three inserts failed for various reasons, which largely came down to the complexity and integrity of the DNA, as well as the size of the fragments.