Team:Cornell/project/wetlab/future work

While we have already made great strides towards improving the accessibility of fungal genetic engineering to industry, academia, and other iGEM teams, we still strive to expand the utility of our toolkit through additional characterization and troubleshooting. In order to apply our work specifically to biomaterials development, we have constructed numerous vectors and are working to perfect a protocol for Agrobacterium tumefaciens-mediated transformation into the Ganoderma lucidum genome [1]. We are also working to incorporate all of our pre-existing fungal expression constructs into Cochliobolus heterostrophus-specific vectors that allow for homologous recombination into the fungal genome. Expression stability will be tested by alternate plating on selective and non-selective plates and maintained by constant selection. Our goal is to test and compare the expression of each of our constructs in Cochliobolus and Ganoderma to provide insight into the expression compatibility of a wide breadth of fungal chassis.

We also plan on acquiring additional fluorescence data in order to quantify the relative strengths of the constitutive promoters in our library on a rating scale, similar to that of the Anderson promoter collection available in the iGEM parts registry [2]. We also specifically hope to use the glucose-repressible, polygalacturonic acid-induced pelA promoter as a controlled, inducible activator of the kill-switch and site-specific recombination systems. Furthermore, we are in the process of characterizing a promoter and terminator from Wageningen University’s iGEM team.

The foundations that we have laid for fungal genetic engineering, and are continually striving to improve, provide a platform for other researchers to apply these tools to a wide variety of sustainable industries.