Team:UNebraska-Lincoln

Welcome to iGEM 2018!

This is University of Nebraska - Lincoln iGEM team

Abstract

The emerald ash borer (EAB) is an invasive species that first appeared in the United States in 2002. As of February 2018, the EAB has spread to 32 states, including Nebraska. The city of Lincoln has 14,000 ash trees that are in danger of EAB infestations within the next few years. Adult EAB lays eggs on the bark of ash trees, and when the larvae hatch, they bore into the trees. They feed on the transportation tissues of the trunk, girdling the tree. Any infested ash tree that is left untreated will die.

It is very important to monitor the EAB population and detect how it is spreading. The information gives governments and organizations more time to create treatment plans and strategies to limit EAB migration. EAB populations are currently monitored by traps baited with green leaf volatiles from ash trees. In the past, traps have also been baited with essential oils from the Manuka and Phoebe trees. These oils are known to contain molecules present in ash bark, but they are no longer used due to inconsistent supplies (McCullough, 2017). Studies have shown that traps baited with both the green leaf volatiles and essential oils are the most effective lures for EAB (Crook, 2010). The six attractive compounds in these oils are α-cubebene, α-copaene, 7-epi-sesquithujene, trans-β-caryophyllene, α-humulene (α-caryophyllene), and eremophilene. Of these, 7-epi-sesquithujene and eremophilene are considered the most important components (Crook, 2010). Neither of these compounds is commercially available, which limits the application and research of their use in traps. Although eremophilene can be synthesized from a commercially available precursor, it is impractical to chemically synthesize 7-epi-sesquithujene in a lab (Crook, 2010).

The goal of the project is to biosynthesize 7-epi-sesquithujene in an efficient and reliable manner. 7-epi-sesquithujene is a sesquiterpene, which is a type of compound known for being difficult to extract and isolate from plants. Research shows that 7-epi-sesquithujene can successfully be synthesized by Escherichia coli through the enzyme 7-epi-sesquithujene synthase (TPS4-B73). This biosynthesis requires the precursor farnesyl pyrophosphate (FPP) (Köllner, 2004), which can be produced in E. coli by engineering the mevalonate pathway found in S. cerevisiae and some enzymes from E. coli itself (Anthony, 2009). By implementing this pathway and the TPS4-B73 gene, E. coli can independently produce desirable levels of synthesis. From there, the 7-epi-sesquithujene can be purified to use in EAB traps.

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