Expanding an algae production platform: Industrially relevant advancement of Chlamydomonas reinhardtii

The initial surge of industrial interest in green algae for the production of renewable fuel has given way to a blossoming industry with the potential to contribute to the commercial production of food, materials, and modern medicine. Algae are biologically diverse, edible, genetically tractable and have high lipid and protein content. Their unicellular nature facilitates the efficient conversion of energy into biological material without the need to produce structural materials like that of higher plants. The diversity of the fields in which algae have become a viable production platform has necessitated development of a broad range of cultivation strategies and tools for improving their industrial relevance. Molecular research on green algae has yielded a growing encyclopedia of genetic tools for manipulation of the chloroplast and nuclear genomes. However, as the landscape of desired algal products has changed, some of these tools need to be refined for new cultivation systems and advanced product engineering. Although the future is bright for algae as a bio-manufacturing platform, many aspects of their production lag behind their established counterparts like yeast, E. coli and mammalian cell culture. The research outlined in this dissertation presents significant gains in the advancement of green algae as an industrial organism by refining cultivation strategies and genetic tools to foster success in commercially relevant systems.Production of algae in both open and closed systems is addressed. For open pond cultivation, environmental concerns render certain genetic markers unusable. A system for selection of transgenic algae without the need for antibiotic resistance cassettes has been developed. In general, closed systems offer more tight regulation of abiotic growth conditions and optimization of growth and product accumulation in these systems is imperative to the future of industrial algae. In closed systems, high density cultures are required for maximum biomass yield with limited infrastructure. Research presented here shows the first look at C. reinhardtii grown in fermentors for high-density cultivation and provides a metabolic comparison with more standard growth conditions. An analysis of transgene expression in high-density cultures expressed a need for new genetic tools that are functional under these conditions. The second half of the dissertation focuses on the development of advanced genetic tools and synthetic systems for optimization of transgene expression. Synthetic tools have the power to revolutionize recombinant protein production in green algae. Ultimately, this dissertation provides an extensive body of work which identifies gaps in the commercial viability of green algae, and provides transformational solutions to many of the problems hampering industrial relevance