Strategies for the selection, identification and improvement of microalgae strains for enhanced biofuel production
AuthorHathwaik, Leyla Trinidy
AdvisorCushman, John C.
Biochemistry and Molecular Biology
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Increasing energy imports, concerns about finite fossil fuel supplies, and a greater recognition of the environmental effects of the combustion of fossil fuels have ignited interest in microalgal-culture for the generation of biofuels. Given their relative high oil and starch content and their fast growth rate, microalgae possess useful attributes as biofuel feedstocks for biofuel production. Microalgae are highly diverse eukaryotic photosynthetic microorganisms consisting of between 200,000 and 800,000 species of which over 40,000 species have been identified. The suitability of microalgae as a biofuel feedstock and the large number of strains emphasizes the need for strategies to effectively select, identify and improve microalgae strains with enhanced lipid or starch production. In order to select algal strains with enhanced traits two different strategies were pursued. The first strategy was based upon continuous Percoll® buoyant density gradient centrifugation (BDGC). Sixty rounds of reiterative, transgressive buoyant density gradient selection resulted in the isolation of cells populations with significantly increased in lipid or starch content. The second approach employed Fluorescent Activated Cell Sorting (FACS) to select and isolate Dunaliella cells containing either high oil or starch amounts based upon fluorescence staining intensities. Thirty rounds of FACS selection resulted in the isolation of cells populations with significantly either increased lipid or starch content. Furthermore, the alterations in feedstock characteristics produced during reiterative selection remained stable after seventy and thirty weeks of selection by BDGC and FACS, respectively. In order to identify various microalgae stains in production settings, molecular markers including the 18S rDNA, photosystem I P700 chlorophyll a apoprotein A2, and the internal transcribed spacer genes were used. In addition, in order to improve microalgae strains for biofuel production a stable plastome transformation method to genetically improve feedstock traits was attempted. Lastly, in order to identify key components in modulating lipid production, changes in mRNA abundance during nutrient starvation, a lipid trigger, were identified by microarray-based mRNA expression profiling. Such information will be invaluable for providing rational strategies for the manipulation of desirable microalgal feedstock characteristics for biofuel production.