Terpenes represent the largest class of natural compounds, which can be used in a wide range of applications from medicines to flavor additives. Current technologies are limited by non-environmentally friendly synthesis techniques (petroleum-based) or inadequate production values (native plant production). To solve this limitation, synthetic biologists have engineered microbes to become terpene production platforms. Although great success has come from the use of some of these platforms, these engineered organisms natively lack high flux through necessary terpene producing pathways. To rectify this problem, extensive metabolic engineering has to be completed in order to obtain industrial-relevant production levels. This dissertation looks to expand the microbial terpene platform idea through the genetic modification of an anaerobic, methane-producing archaeon whose metabolic substrates and byproducts may allow for improved terpene production than previously used microbial models. Methanosarcina is a genus of methanogens that natively produces high levels of prenyl precursors for cellular membrane synthesis, leading to the hypothesis that this pathway can be tapped for terpene production. To show the genus ability to produce high levels of a desired terpene, a modified genetic system was employed to genetically engineer Methanosarcina acetivorans C2A and Methanosarcina barkeri Fusaro to express a functional isoprene synthase. Genetic modification of these strains by the addition of a single enzyme involved in isoprene biosynthesis results in high levels of isoprene production with negligible impact on microbial fitness. The isoprene produced is at levels comparable with existing isoprene producing microbes; but an advantage of Methanosarcina use is in feedstock as it can be readily grown on cheap or often free, waste products. To further the idea that Methanosarcina can be developed into a terpene production platform, strains overexpressing isoprenyl diphosphates synthase were created. Such strains showed increased lipid production compared to wild-type, indicating that high carbon flux is flowing through the terpene precursor pathway. This leads to the promising conclusion that Methanosarcina can be used as the desired platform system
