Team:Cornell/project/wetlab/fungal toolkit

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<b>Scroll over each icon to find out more about the functional requirements for our device!</b>
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<b>Scroll over each icon to find out more about the components of our toolkit!</b>
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<h5>Inlet</h5>
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Water flows into our system, providing a sample to test water quality.
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<h5>Regulatory Elements</h5>
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Provide modular protein expression controls for genetic constructs.
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<h5>Filter</h5>
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Prevents foreign microbes from entering and contaminating our reactor.
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<h5>Selectable Markers</h5>
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Allow for the selection of desired fungal transformants.
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<h5>Pumps</h5>
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Move water and food to and from the reactor.
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<h5>Characterization</h5>
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Quantifies gene expression levels using fluorescent proteins.
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<h5>Piping and Calibration</h5>
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Resist corrosion and set flow rates to the optimal level.
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<h5>Homologous Constructs</h5>
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Enable integration of genetic constructs into specific regions of fungal genomes.
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<h5>Reactors</h5>
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House bacteria that produce electrical current in response to toxins.
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<h5>Biosafety</h5>
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Implements measures to prevent horizontal gene transfer and induce a kill switch.
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<h5>Continuous</h5>
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Pollution events can be hard to spot, making discrete testing inadequate as well as expensive. Monitoring water quality is a 24 hour job.
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<h5>Field Deployable</h5>
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Industrial water monitoring is needed in remote and rugged terrain. In order to be applicable in these terrains, our device should be durable, water-proof. Being out in the field also means all food for bacteria and power for electronics must be provided by the device itself.
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To collect data in isolated locations, wireless communication is essential. This requires digital conversion of signals from our bacteria, so that it can be transmitted to the user and accessed online.
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To monitor water quality in a truly continuous fashion, our device must be able to sustain itself without maintenance for long periods of time. Frequent maintenance is impractical.
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<h3>Protoplasting</h3>
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<h5>Background</h5>
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Transformation of plant and fungal cells is difficult due to their cell walls that block passage of foreign DNA. Protoplasting is the method by which the cells walls of plant and fungal cells are digested to produce cells without cell walls, called protoplasts. Through additional methods, such as electroporation and PEG transformation, DNA can be uptaken by the protoplasts and then regrown into cells containing specific genes.
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<h5>Method</h5>Usually this species is protoplasted using lywallzyme and novozyme. However, these enzymes are exclusively available in China. So, protoplasting was attempted with driselease and glucanex instead. After several attempts, we found that the enzymes are unable to digest the cell wall without also killing the cell. In addition, the Ganoderma mycelium was hard to pellet when centrifuging and made the protoplasting procedure difficult. Thus, <i>Cochliobolus heterostrophus</i> was used instead. Protoplasting was successful using driselease and glucanex and then transformed via PEG solution.   
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<h3>References</h3>
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Sun L, Cai H, Xu W, Hu Y, Gao Y, Lin Z (2001). Efficient Transformation of the Medicinal Mushroom Ganoderma lucidum. Plant Molecular Biology Reporter, 19, 383a-383j.
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Amir Sharon (ed.), <i>Molecular and Cell Biology Methods for Fungi</i>, Methods in Molecular Biology, vol. 638. Turgeon BG, Condon B, Liu J, Zhang N (2010). Protoplast Transformation of Filamentous Fungi.
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Latest revision as of 13:01, 12 October 2013

Cornell University Genetically Engineered Machines

Fungal Toolkit

Scroll over each icon to find out more about the components of our toolkit!
Regulatory Elements
Provide modular protein expression controls for genetic constructs.
Selectable Markers
Allow for the selection of desired fungal transformants.
Characterization
Quantifies gene expression levels using fluorescent proteins.
Homologous Constructs
Enable integration of genetic constructs into specific regions of fungal genomes.
Biosafety
Implements measures to prevent horizontal gene transfer and induce a kill switch.

Retrieved from "http://2013.igem.org/Team:Cornell/project/wetlab/fungal_toolkit"