Enhancing lipid production of Lipomyces starkeyi for the upgrading of C6 sugars from lignocellulosic biomass

Lipomyces starkeyi is capable of producing greater than 60% of its mass as lipids and is currently a leading candidate for strain development for the production of fatty acids as fuel precursors using a C6 sugar rich stream from lignocellulosic biomass.  In addition to its high lipid productivity, its genome sequence is available and genetic tools for transformation have already been reported, however significant challenges remain in engineering its genome. Specifically, targeted gene integration or knockout have not yet been developed making directed engineering for enhanced lipid production currently very challenging.  As such, there is a need to develop genetic tools in L. starkeyi necessary for efficient strain manipulations (transformation optimization, enhancing homologous recombination, marker-recycling system for integration, mutagenesis, etc.) to accelerate genetic manipulations in the future.  Here, we describe some of our efforts to make L. starkeyi genetically tractable.  In addition to modifying particular gene targets to increase our capacity to perform genetic engineering in L. starkeyi, we are also exploring classical strain improvement techniques.  Specifically, we are using random mutagenesis coupled with selection using fatty acid synthase inhibitors as a mechanism for improving lipid production. We have obtained several candidate isolates, which show improved growth in the presence of fatty acid synthase inhibitors and were evaluated for lipid production.