Genetically Engineered \u3ci\u3eThermosynechococcus Elongatus\u3c/i\u3e BP1: Assessment of Potential Biorisks and Biofuel Production

According to the International Energy Outlook 2019, released by the U.S. Energy Information Administration, it is projected that the energy consumption will increase up to 50% between 2018 and 2050 worldwide. As fossil fuel being a finite source of energy with the risk of depletion, many countries are now facing an energy security crisis. Therefore, it is important to develop other renewable and sustainable energy sources that will allow countries to shift away from depending on fossil fuels. Among several types of renewable energy, biofuel production using genetically engineered cyanobacteria is capturing much interest due to its many advantages. Different forms of biofuels such as ethanol, butanol, isobutanol, biodiesel have been successfully produced using cyanobacteria. Though this is a promising approach in achieving a renewable and sustainable energy source, research is still in its early stage and there are multiple aspects yet to be done. Here, the first aim of this work seeks to answer the question of whether genetically engineered cyanobacteria could pose a biosafety risk by transferring their gene(s) into other bacteria upon contact. This was done via a horizontal gene transfer study which included co-culturing of genetically engineered cyanobacteria carrying kanamycin resistant gene and wild-type E. coli DH5α. E. coli cells were then screened for kanamycin resistant gene after being in the same environment with the genetically engineered cyanobacteria. By doing so, it would provide a better understanding of the risk this approach might have. In addition, this work discussed the use of a thermophilic strain of cyanobacteria, Thermosynechococcus elongatus BP1, for isobutanol production. It is expected that the thermophilic property of this strain would provide some biosafety features which could eliminate some environmental as well as ecological risks. A cassette carrying a set of genes is inserted into Thermosynechococcus elongatus BP1 cells and isobutanol production was detected and quantified by GC-MS. Lastly, this dissertation also investigated the tolerance of Thermosynechococcus elongatus BP1 toward different alcohols by supplementing cell cultures with different concentrations of ethanol, isobutanol, and 1-butanol to help understand the effect of these alcohols have on cell growth for industrial scaled up