Team:Peking/Project

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<h3 class="title1">Synthetic Biology: What's Hindering?</h3>
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<h3 id="title1">Synthetic Biology: What's Hindering Us?</h3>
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<p>Viewing cells as programmable entities, efforts in synthetic biology have focused on the creation and perfection of biological modules and systems in order to achieve specific functions. Since first demonstrated by the bacterial toggle switch and the repressilator in 2000, synthetic biologists now own the ability to engineer genetic circuits, metabolic pathways, and even genomes. <br/><br/>During genetic engineering, chemicals are frequently employed as signals to control molecular and cellular behavior. However, because chemicals deliver information only by diffusion (Figure 1), chemical signals are seriously limited by the short-range interaction. Another serious issue regarding chemical signals is the difficulty to erase them, which makes it difficult for the system to reset. Additionally, the cytotoxicity, high cost and narrow dynamic range of chemicals urge synthetic biologists to find more spatiotemporally specific and environmental friendly alternatives.</p>
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<p>Efforts in synthetic biology have been primarily focused on the creation and perfection of biological modules and systems in order to achieve specific functions. Since first demonstrated by the bacterial toggle switch and the Repressilator in 2000, synthetic biologists now possess the ability to engineer and manipulate genetic circuits, metabolic pathways, and even entire genomes. <BR><BR>
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In genetic engineering, chemicals are frequently employed as signals to control molecular and cellular behavior. However, because chemicals deliver information only by diffusion, chemical signals are seriously limited by the short-range interactions. Another serious issue regarding chemical signals is the complexity of the process in fully eradicating them, making it difficult for the system to reset. Additionally, the cytotoxicity, high cost, and narrow dynamic range of chemicals urge synthetic biologists to find more spatiotemporally specific and environmental friendly alternatives.
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<img src="/wiki/images/b/be/Peking2012_Chemical_Light_comparison.png" alt="comparison of Chemical and light" style="width:600px"/>
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  <p class="description">Figure 1. The diffusion of chemicals.</p>
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<p class="description" style="text-align:center;font-style:normal;">Figure 1. Comparison of chemical signal and light signal</p>
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<h3 class="title2">Optogenetics: Inspire Synthetic Biology </h3>
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<p>During the last decade, optogenetics has made significant impacts on life sciences and other practices. Literally, optogenetics refers to the genetic strategies which employ light as the input signal in the place of chemical signals. The spatiotemporal specificity of light signals allows for precise manipulation and long range interaction without physical contact. Besides, light resources are cheaper, more sustainable and environmentally friendly. <br/><br/>Due to the advantages mentioned above, optogenetics has evolved rapidly: light-switchable promoters, light-induced protein-protein interaction, light-activated ion channels and eventually, light-controlled animal behavior (Figure 2).  
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<p>Literally, optogenetics refers to the genetic strategies which employ light as the input signal in the place of chemical signals. During the last decade, optogenetics has made significant impact on life sciences and other practices<sup><a href="#ref1" title=" Miesenbock, G. (2011). Optogenetic control of cells and circuits." Annu Rev Cell Dev Biol 27: 731-758.>[1]</a></sup>. It has evolved rapidly, with light-switchable promoters<sup><a href="#ref2" title=" Shimizu-Sato, S., E. Huq, et al. (2002). A light-switchable gene promoter system". Nat Biotechnol 20(10): 1041-1044.>[2]</a></sup>, light-induced protein-protein interaction<sup><a href="#ref3" title=" Levskaya, A., O. D. Weiner, et al. (2009). Spatiotemporal control of cell signalling using a light-switchable protein interaction." Nature 461(7266): 997-1001.>[3]</a></sup>, light-activated ion channelsand<sup><a href="#ref4" title=" Zemelman, B. V., G. A. Lee, et al. (2002). Selective photostimulation of genetically chARGed neurons". Neuron 33(1): 15-22.>[4]</a></sup>  eventually, light-controlled animal behavior, and so on. The spatiotemporal specificity of light signals allows for precise manipulation and long range interaction without physical contact. Besides its incredible range of functions, light resources are also cheaper, more sustainable, and environmentally friendly. These properties together make optogenetic tools attractive for synthetic biologists.
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Figure 2. Revolution of Optogenetics.</p>
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Figure 2. Evolution of Optogenetics<sup><a href="#ref6">[6]</a><sup></p>
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<p>Light as a signal may carry massive information. The wavelength, intensity, frequency and spatial-temporal patterns of light are all informations encoded in a physical manner. Plus light signals possess a Bool-logic-like dynamic behavior and does not require any medium for its propagation, it would be much more enabling in transmitting accurate and complex information than the frequently investigated chemical signals. (In fact, natural biological systems has already explored this advantage to an astonishing extent. Think of our vision! ) <br/><br/>
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Thus using light as a signal, we may program cellular behaviors to achieve, say, cell-cell communication that does not require physical contact. To communicate through light, first cells must be able to sense light, which means we need to develop a light sensor. And to realize this new generation of light communication that has never been implemented before, the sensor itself must make a big difference.</p>
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<p>However, current optogenetic tools are far from satisfactory for synthetic biology application. In next section, we will comprehensively analyze the current optogenetic methods and raised issues that our <i>Luminesensor</i> needs to circumvented.
 
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<h3 class="title3">Potential Problems of Current Optogenetic<br/>Methods</h3>
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In order to evaluate the current optogenetics methods in a systematic manner, this year’s Peking iGEM team has carefully selected 15 optogenetics methods, ranging from applications in prokaryotes to potential uses in eukaryotes. By statistically evaluating the modularity, sensitivity, and dynamic range of these optogenetics methods, we have come to a conclusion to address four potential problems of current optogenetics approaches:
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1. Miesenbock, G. (2011) Optogenetic control of cells and circuits. <i>Annu. Rev. Cell. Dev. Biol.</i>, 27: 731: 758<br />
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<p>1. Low sensitivity<br/>
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2. Shimizu-Sato, S., Huq, E., <i>et al</i>. (2002) A light-switchable gene promoter system. <i>Nat. Biotechnol.</i>, 20(10): 1041: 1044<br />
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To minimize the influence on cellular metabolism, strong light needs to be avoided for practical applications. Almost 50% of the 15 optogenetics tools employ the use of lasers as the light source. We demonstrated the effect of lasers on the organisms by illuminating a lawn of <i>E.coli</i> with a laser pointer. As clearly shown in Figure 3, no bacterial growth could be found on the spot that was illuminated. Only about 25% of the 15 optogenetics tools are able to sense natural light (~1W/m<sup>2</sup>), an obviously cheaper and much more sustainable source of light (Figure 4).
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3. Levskaya, A., Weiner, O.D., <i>et al.</i> (2009) Spatiotemporal control of cell signalling using a light-switchable protein interaction. <i>Nature</i>, 461(7266): 997: 1001.<br />
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4. Zemelman, B.V., Lee, G.A., <i>et al.</i> (2002) Selective photostimulation of genetically chARGed neurons. <i>Neuron</i>, 33(1): 15: 22<br />
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  Figure 3. The effect of lasers on bacterial growth.</p>
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5. Ohlendorf, R., Vidavski, R.R., <i>et al.</i> (2012) From dusk till dawn: one-plasmid systems for light-regulated gene expression. <i>J. Mol. Biol.</i> 416(4): 534: 542<br />
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6. http://www.stanford.edu/group/dlab/optogenetics/
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  Figure 4. The sensitivity of the 15 optogeneticsmethods.</p>
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<p>Narrow dynamic range<br/>Many cross-species optogenetics methods require the addition of exogenous chromophores, e.g. phycocyanobilin (PCB) and caged amino acids. This leads to two major problems: (1) the exogenous chromophores may be incompatible with the endogenous components, and observed in some cases, may even have some influence on the bacterial growth; (2) the inhomogeneity caused by the diffusion of chromophores, an undesired AND gate, is constructed in these methods (Figure 6), which makes the output unpredictable.  
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Figure 5. The dynamic range of the 15 optogenetics methods.</p>
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<p>Dependency on exogenous chromophores<br/>Many cross-species optogenetics methods require the addition of exogenous chromophores, e.g. phycocyanobilin (PCB) and caged amino acids. This leads to two major problems: (1) the exogenous chromophores may be incompatible with the endogenous components, and observed in some cases, may even have some influence on the bacterial growth; (2) the inhomogeneity caused by the diffusion of chromophores, an undesired AND gate, is constructed in these methods (Figure 6), which makes the output unpredictable.
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Figure 2. Both chromophores AND light are required to activate/inactivate the system.</p>
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<p>Limited application in prokaryotes<br/>
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Another serious issue concerns the limited application of optogenetics methods in prokaryotes. The rapidly reproducing prokaryotes play a very significant role in both fundamental research and industrial application. In many methods, only proof of concept is provided, without practical application concerned. The low sensitivity, narrow dynamic range,   and need for exogenous chromophores, as mentioned above, are the main blocks of a broader and more practical application of these methods.
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<h3 class="title4">A New Generation of Optogenetics</h3>
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By gaining insight into the current optogenetics methods, the Peking iGEM team speculates that rationally designing a robust and ultrasensitive transcription factor may circumvent all the potential problems mentioned above: high sensitivity allows dimmer, cheaper and safer light as the source of signals; because the light sensitive transcription factor functions directly on the gene expression, the basal level may be low enough to allow a high dynamic range. <br/><br/>Now that such an ultrasensitive and efficient optogenetics module has been successfully designed, the Peking iGEM endeavors to explore the full proficiency of optogenetics and raise a new generation of optogenetics with broader applications on the cellular behavior level, the intercellular communication level, and the industrial level.
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Latest revision as of 09:31, 26 October 2012

Synthetic Biology: What's Hindering Us?

Efforts in synthetic biology have been primarily focused on the creation and perfection of biological modules and systems in order to achieve specific functions. Since first demonstrated by the bacterial toggle switch and the Repressilator in 2000, synthetic biologists now possess the ability to engineer and manipulate genetic circuits, metabolic pathways, and even entire genomes.

In genetic engineering, chemicals are frequently employed as signals to control molecular and cellular behavior. However, because chemicals deliver information only by diffusion, chemical signals are seriously limited by the short-range interactions. Another serious issue regarding chemical signals is the complexity of the process in fully eradicating them, making it difficult for the system to reset. Additionally, the cytotoxicity, high cost, and narrow dynamic range of chemicals urge synthetic biologists to find more spatiotemporally specific and environmental friendly alternatives.

comparison of Chemical and light

Figure 1. Comparison of chemical signal and light signal

Optogenetics: Inspiring Synthetic Biology

Literally, optogenetics refers to the genetic strategies which employ light as the input signal in the place of chemical signals. During the last decade, optogenetics has made significant impact on life sciences and other practices[1]. It has evolved rapidly, with light-switchable promoters[2], light-induced protein-protein interaction[3], light-activated ion channelsand[4] eventually, light-controlled animal behavior, and so on. The spatiotemporal specificity of light signals allows for precise manipulation and long range interaction without physical contact. Besides its incredible range of functions, light resources are also cheaper, more sustainable, and environmentally friendly. These properties together make optogenetic tools attractive for synthetic biologists.

[Fig 2.]

Figure 2. Evolution of Optogenetics[6]

Light as a signal may carry massive information. The wavelength, intensity, frequency and spatial-temporal patterns of light are all informations encoded in a physical manner. Plus light signals possess a Bool-logic-like dynamic behavior and does not require any medium for its propagation, it would be much more enabling in transmitting accurate and complex information than the frequently investigated chemical signals. (In fact, natural biological systems has already explored this advantage to an astonishing extent. Think of our vision! )

Thus using light as a signal, we may program cellular behaviors to achieve, say, cell-cell communication that does not require physical contact. To communicate through light, first cells must be able to sense light, which means we need to develop a light sensor. And to realize this new generation of light communication that has never been implemented before, the sensor itself must make a big difference.

Reference

  • 1. Miesenbock, G. (2011) Optogenetic control of cells and circuits. Annu. Rev. Cell. Dev. Biol., 27: 731: 758
  • 2. Shimizu-Sato, S., Huq, E., et al. (2002) A light-switchable gene promoter system. Nat. Biotechnol., 20(10): 1041: 1044
  • 3. Levskaya, A., Weiner, O.D., et al. (2009) Spatiotemporal control of cell signalling using a light-switchable protein interaction. Nature, 461(7266): 997: 1001.
  • 4. Zemelman, B.V., Lee, G.A., et al. (2002) Selective photostimulation of genetically chARGed neurons. Neuron, 33(1): 15: 22
  • 5. Ohlendorf, R., Vidavski, R.R., et al. (2012) From dusk till dawn: one-plasmid systems for light-regulated gene expression. J. Mol. Biol. 416(4): 534: 542
  • 6. http://www.stanford.edu/group/dlab/optogenetics/
  • Totop Totop

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