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Browsing by Subject "Yarrowia lipolytica"
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Item ENGINEERING AN OLEOGINOUS YEAST FOR THE PRODUCTION OF BIODIESEL(Office of the Vice Chancellor for Research, 2012-04-13) Fernandez, Astroski, J.; Lewis, P.; McDaniel, J.; Blacklock, B.J.; Minto, R.E.; Randall, S.K.There are economic and social interests in replacing the current energy dependence we have on petroleum-based oleochemicals. Yarrowia lipolytica, an oleaginous yeast, has the ability to metabolize unique carbon sources, particularly hydrocarbons and to accumulate large amounts of lipids which could be developed into a source of biodiesel. The ability of Y. lipolytica to accumulate triacylglycerols in lipid droplets and the complete sequencing of its genome make Y. lipolytica a viable organism to genetically engineer for the production of large quantities of biodiesel precursors. The purpose of this project is to genetically modify Y. lipolytica to further increase its production of triacylglycerols by knocking out genes that encode enzymes involved in the β-oxidation of fatty acids. This genetic modification will be accomplished by using homologous recombination to disrupt the genes POX3-5 and POT1. The 5’ and 3’ untranslated regions of POX3-5 and POT1 were amplified by polymerase chain reaction and cloned to allow a drug resistance gene to be introduced between them. Following cloning, these genes will be knocked out from the Y. lipolytica genome using drug resistance as a marker. The disruption of these genes is expected to increase the accumulation of triacylglycerols in Y. lipolytica lipid droplets versus the wild-type. Progress towards the goals of this project will be reported.Item Investigations of lipid metabolism in Yarrowia lipolytica(2014-07-31) Blocher-Smith, Ethan Charles; Minto, Robert; Long, Eric C. (Eric Charles); Jones, LisaAn investigation of the lipid metabolism pathway in the yeast Yarrowia lipolytica was conducted. Yarrowia is an oleaginous ascomycete that is capable of growing on many different substrates, which derives its name from its high efficiency of growth on lipids. Once the exogenous lipids are converted into free fatty acids and internalized by the yeast, the primary mode of degradation is through β-oxidation mediated by the peroxisomal oxidases, or POX genes. These enzymes catalyze the formation of a trans double bond, producing the trans-2-enoyl product. Our study looked at the comparison of the Y. lipolytica prototrophic strain against a knockout of the Pox2 gene on the uptake, incorporation, and degradation of relevant fatty acids. To construct this gene knockout, a novel gene deletion method using a combination of Cre recombinase and the AHAS* gene was synthesized, developed, and tested successfully. This knockout system allows for serial deletion of genes with the use of only one resistance marker, with excision of the marker after selection.