Abstract:
The model eukaryotic green microalga Chlamydomonas reinhardtii has been shown to be an interesting platform organism for biotechnological application and production of isoprenoid natural products through metabolic engineering. C. reinhardtii uses light for energy and carbon dioxide (CO2) as its main carbon source for growth. The work presented in this thesis modified a genetically tractable C. reinhardtii strain (UVM4) to create a strain called UPN22, which can grow on phosphite as a sole P source and Nitrate as a sole N source through plastid and nuclear genome engineering, respectively. These nutrient sources can help minimize contamination and maximize cell densities in photoautotrophic cultivations. The capacity of the engineered strain to express a range of recombinant proteins from its chloroplast genome as an alternative application of this strain was investigated. No reliable results were obtained, and further genetic optimization for chloroplast expression of transgenes is suggested. UPN22 was further engineered to produce the heterologous C15 sesquiterpenoid patchoulol by transforming the nuclear genome with constructs encoding a sesquiterpene synthase. This synthase was targeted to the cytoplasm, algal plastid, or both, to investigate the capacity of the cell to produce heterologous sesquiterpenoid in different sub-cellular compartments. This was conducted in strains with and without knockdown of the native squalene synthase pathway to determine the dynamics of sesquiterpenoids biosynthesis when squalene was perturbed. Additionally, the constructs were modified by fusion with yeast farnesyl pyrophosphate synthase (FPPS) to enhance the local production of farnesyl pyrophosphate (FPP) pools in each context. This work will help guide future efforts for engineering and use of C. reinhardtii for sesquiterpenoid bio-process concepts.

Bio:
PhD candidate in Bioengineering at King Abdullah University of Science & Technology (KAUST) with a Master’s degree in Cell and Molecular Biology from KAUST as well, and a Bachelor’s in Genome and Biotechnology from King Abdul-Aziz University. Strong foundation in bioengineering and molecular biology.