Team:Tokyo Tech/Result

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!align="center"|[[Team:Tokyo_Tech/Result|Result]]
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!align="center"|[[Team:Tokyo_Tech/Acknowledgements|Acknowledgements]]
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== Protocol ==
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== <font size=5> Checking pressure response ability of Plac </font> ==
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[[Image:Transfer_Function.png|thumb|300px|left|figure 1 Hill function]]
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=== IPTG assay===
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'''Objective'''
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<p class=MsoNormal>We examined changing of GFP fluorescence strength with pressurizing.At the same time, we examined fluorescence strength by adding IPTG under 0.1MPa.As a way of observation, we adopted a fluorescence microscope, FACS and FLA.FACS and FLA is for quantitative analysis of fluorescence strength. FLA’s result is Fig2, Fig3, Fig4.</font></p>
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<p class=MsoNormal>Fig 2: Average of Plac /p></td>
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<td class="a" width="40%">　Fig 3: Average of ⊿p </td>
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<p class=MsoNormal>Fig 2: Average of Plac /p></td>
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<td class="a" width="40%">　Fig 3: Average of ⊿p </td>
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By testing how LacI represses the lac promoter, Hill coefficient of lac promoter should be decided. In order to adjust effective concentration of LacI, IPTG was added.
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          <td class="a">　Fig 4: Average of Ptet</td>
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          <td class="a" width="40%"> Fig 5: Fig2&3&4 gathers into one chart.</td>
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<p class=MsoNormal>Figure 2, 3, 4 ; These charts show change of Geo Mean under each pressures. Figure 2 is about Plac-GFP. We constructed Plac-GFP. E. coli strains is JM109. Between 0.1MPa and 30MPa, Geo Mean almost don’t change.But at 50MPa Geo Mean increase sharply. Figure 3 is about promoter less-GFP (∆p-GFP) as a negative control. Between 0.1MPa and 30MPa, Geo Mean almost don’t change. But, at 50MPa Geo Mean increase sharply. Figure 4, is about Ptet-GFP as a positive control.Increasing pressure, Geo Mean increases. </p></td>
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'''Result & Conclusion'''
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A fluorescence microscope is for qualitative analysis of fluorescence strength. Fluorescence microscope image is Fig 6.</td>
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GFP fluorescence intensity was enhanced in an IPTG-dose dependent manner. It indicates that the LacI repression was getting weak by adding IPTG. Taken together with the graph shown in figure 1 and the formulation of Hill function fitting described above, the characteristics of the lac promoter expressed in Hill function was determined.
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Latest revision as of 13:49, 26 October 2008

Home Construction Acrylic container Development of promoter Genetic toggle switch Parts Submitted to the Registry Result The Team Acknowledgements

Protocol

figure 1 Hill function

IPTG assay

Objective

By testing how LacI represses the lac promoter, Hill coefficient of lac promoter should be decided. In order to adjust effective concentration of LacI, IPTG was added.

Result & Conclusion

GFP fluorescence intensity was enhanced in an IPTG-dose dependent manner. It indicates that the LacI repression was getting weak by adding IPTG. Taken together with the graph shown in figure 1 and the formulation of Hill function fitting described above, the characteristics of the lac promoter expressed in Hill function was determined.

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