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Team:Cornell/project/wetlab
From 2013.igem.org
Wetlab Overview
We are working with the fungus Ganoderma lucidum, a model organism for the class of wood-rot fungi that is used in Ecovative’s products. We have assembled a number of constructs for the production of carotenoid pigmentation, fluorescent proteins, as well as simple antibiotic resistances that we are seeking to transform by both random insertion and homologous recombination into the organism’s genome. These proof-of-concept tools are necessary to optimize our process so that we can then obtain rigorous, reliable characterization data for the antifungal gene we are seeking to introduce, a protein that specifically targets Aspergillus niger.
However, as Ganoderma does not have a well-standardized transformation protocol and takes several days to grow to an appropriate density, we are also conducting much of our basic fungal characterization work in Cochliobolus heterostrophus, a simpler fungus. In addition, we are seeking to introduce a novel T7 viral regulation system into fungi; similar systems have been used successfully in mammalian cells, and this system would greatly expand the accessibility of fungal genetic engineering beyond experienced mycologists. This system allows us to conduct preliminary characterization within E. coli BL21-A1, a much simpler system for acquiring quantitative data.
We are also implementing a number of biosafety measures within our toolkit, including a recombination system that has previously been demonstrated to work in simpler fungi, which will allow us to remove antibiotic resistance genes and targeted antifungal compounds before the end of the production process, in order to prevent them from spreading into the environment. The strain we are seeking to develop would be a huge boon to the economic viability of Ecovative’s sustainable biomaterials, as it would allow them to more easily maintain the high levels of quality control that are necessary in scaled-up production.
However, as Ganoderma does not have a well-standardized transformation protocol and takes several days to grow to an appropriate density, we are also conducting much of our basic fungal characterization work in Cochliobolus heterostrophus, a simpler fungus. In addition, we are seeking to introduce a novel T7 viral regulation system into fungi; similar systems have been used successfully in mammalian cells, and this system would greatly expand the accessibility of fungal genetic engineering beyond experienced mycologists. This system allows us to conduct preliminary characterization within E. coli BL21-A1, a much simpler system for acquiring quantitative data.
We are also implementing a number of biosafety measures within our toolkit, including a recombination system that has previously been demonstrated to work in simpler fungi, which will allow us to remove antibiotic resistance genes and targeted antifungal compounds before the end of the production process, in order to prevent them from spreading into the environment. The strain we are seeking to develop would be a huge boon to the economic viability of Ecovative’s sustainable biomaterials, as it would allow them to more easily maintain the high levels of quality control that are necessary in scaled-up production.
