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Team:Valencia/WetLab/YeastTeam
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| - | At WetLab we are working with '''yeasts''' in order to create each and everyone of the cell-pixels of our bioscreen. Since we want to make yeasts shine, we are using an aequorin-transformed yeast strains. | + | <!--At WetLab we are working with '''yeasts''' in order to create each and everyone of the cell-pixels of our bioscreen. Since we want to make yeasts shine, we are using an aequorin-transformed yeast strains. |
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| - | To make light, Aequorin uses '''coelenterazine''' as its cofactor. Aequorin also needs the binding of Ca<SUP>2+</SUP> to produce light. You can see the complete reaction in the next picture: | + | To make light, Aequorin uses '''coelenterazine''' as its cofactor. Aequorin also needs the binding of Ca<SUP>2+</SUP> to produce light. You can see the complete reaction in the next picture:--> |
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| + | The project our team imagined is about controlling electrically genetically engineered cells to induce a desired behavior. In particular, our Project had the ambitious goal of making LECs (Light Emitting Cells). | ||
| + | In order to do so, we have worked with a bioluminiscent protein named aequorin. Aequorin is present in jellyfish and it is responsible of the emission of Light by these marine invertebrates. In order for aequorin to produce Light, this protein has to be coupled to a prostetic group: coelenterazine. Coelenterazine is oxidized when calcium ions bind to the aequorin-coelenterazine complex and, as a result, light is emitted. This is not a fluorescent reaction, as aequorin do not need to be excited with UV light. | ||
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| + | But since we did not want to work with jellyfishes in the lab, we used budding yeast, Saccharomyces cerevisiae, as a simple host for our experiments. We used a genetically modified yeast strain expressing aequorin. This strain has been reported to produce light through a calcium channel-based system in response to a chemical stimulation. | ||
| - | + | But our aim was to control electrically our LECs, so we tried to induce a membrane depolarization by supplying electricity to our yeasts and open voltage-dependent calcium channels to produce a calcium entry to cytosol and, as a result, get light emission. | |
| - | + | <!--<b>[https://2009.igem.org/Image:Aquorin_val.JPG Aequorin]</b> + <b>[https://2009.igem.org/Image:Coelenterazine.png Coelenterazine]</b> + Ca<SUP>2+</SUP> --> [[Image:Aquorin in action.jpg|350px]]--> | |
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| + | <!--We will use a '''chemical input''' like KOH (alkali shock) to open '''Ca<SUP>2+</SUP> channels''' in the yeasts' membrane (Viladevall L, et al. J Biol Chem. (2004) 279 43614–43624), then Ca<SUP>2+</SUP> will enter into the cells and we will get '''light''' as an '''output'''. We have a [https://2009.igem.org/Team:Valencia/WetLab/YeastTeam/Protocols protocol] and diferent [https://2009.igem.org/Team:Valencia/WetLab/YeastTeam/Experimental experimental designs] to try to reproduce the Arinyo's experiment. | ||
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| + | If we succed in this, we will try to reproduce the same response with an '''electrical stimulus''', using some hardware built by ourselves. We will use an electrical input to open the Ca<SUP>2+</SUP> channels. We hope that we see light by this metod.--> | ||
[[Image:Wiki.JPG|400px|center]] | [[Image:Wiki.JPG|400px|center]] | ||
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| + | In summary, we designed a yeast-based system for the production of light through a jellyfish protein sensitive to calcium. Since calcium enters the cytoplasm through voltage channels, it is theoretically possible to have light emission under electrical control. If this was possible, a biological display made of aequorin-expressing yeasts as living pixels could be constructed. | ||
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Revision as of 17:50, 21 October 2009
WetLab Overview
The project our team imagined is about controlling electrically genetically engineered cells to induce a desired behavior. In particular, our Project had the ambitious goal of making LECs (Light Emitting Cells). In order to do so, we have worked with a bioluminiscent protein named aequorin. Aequorin is present in jellyfish and it is responsible of the emission of Light by these marine invertebrates. In order for aequorin to produce Light, this protein has to be coupled to a prostetic group: coelenterazine. Coelenterazine is oxidized when calcium ions bind to the aequorin-coelenterazine complex and, as a result, light is emitted. This is not a fluorescent reaction, as aequorin do not need to be excited with UV light.
But since we did not want to work with jellyfishes in the lab, we used budding yeast, Saccharomyces cerevisiae, as a simple host for our experiments. We used a genetically modified yeast strain expressing aequorin. This strain has been reported to produce light through a calcium channel-based system in response to a chemical stimulation.
But our aim was to control electrically our LECs, so we tried to induce a membrane depolarization by supplying electricity to our yeasts and open voltage-dependent calcium channels to produce a calcium entry to cytosol and, as a result, get light emission.
-->
In summary, we designed a yeast-based system for the production of light through a jellyfish protein sensitive to calcium. Since calcium enters the cytoplasm through voltage channels, it is theoretically possible to have light emission under electrical control. If this was possible, a biological display made of aequorin-expressing yeasts as living pixels could be constructed.
