iGEM 2010 — Team Imperial_College_London

[![](https://2010.igem.org/images/wiki.png)"](/Main_Page "Main Page")

* [Page](/Team%3AImperial_College_London/Notebook)
* [Discussion](/wiki/index.php?title=Talk:Team:Imperial_College_London/Notebook&action=edit&redlink=1)
* [View source](/wiki/index.php?title=Team:Imperial_College_London/Notebook&action=edit)
* [History](/wiki/index.php?title=Team:Imperial_College_London/Notebook&action=history)
* teams

* [Log in](/wiki/index.php?title=Special:UserLogin&returnto=Team:Imperial_College_London/Notebook "You are encouraged to log in; however, it is not mandatory [o]")

# Team:Imperial College London/Notebook

### From 2010.igem.org

This is a template page. READ THESE INSTRUCTIONS.

You are provided with this team page template with which to start the iGEM season. You may choose to personalize it to fit your team but keep the same "look." Or you may choose to take your team wiki to a different level and design your own wiki. You can find some examples [HERE](https://2008.igem.org/Help%3ATemplate/Examples).

You **MUST** have a team description page, a project abstract, a complete project description, a lab notebook, and a safety page. PLEASE keep all of your pages within your teams namespace.

|  |  |
| --- | --- |
| You can write a background of your team here. Give us a background of your team, the members, etc. Or tell us more about something of your choosing. | [File:Imperial College London logo.png](/File%3AImperial_College_London_logo.png "File:Imperial College London logo.png") 200px |
| *Tell us more about your project. Give us background. Use this is the abstract of your project. Be descriptive but concise (1-2 paragraphs)* | [File:Imperial College London team.png](/File%3AImperial_College_London_team.png "File:Imperial College London team.png") Your team picture |
|  | [Team Example](/Team%3AImperial_College_London "Team:Imperial College London") |

| [Home](/Team%3AImperial_College_London "Team:Imperial College London") | [Team](/Team%3AImperial_College_London/Team "Team:Imperial College London/Team") | [Official Team Profile](https://igem.org/Team.cgi?year=2010&team_name=Imperial_College_London) | [Project](/Team%3AImperial_College_London/Project "Team:Imperial College London/Project") | [Parts Submitted to the Registry](/Team%3AImperial_College_London/Parts "Team:Imperial College London/Parts") | [Modeling](/Team%3AImperial_College_London/Modeling "Team:Imperial College London/Modeling") | **Notebook** | [Safety](/Team%3AImperial_College_London/Safety "Team:Imperial College London/Safety") |
| --- | --- | --- | --- | --- | --- | --- | --- |

## Notebook

You should make use of the calendar feature on the wiki and start a lab notebook. This may be looked at by the judges to see how your work progressed throughout the summer. It is a very useful organizational tool as well.

Retrieved from "[http://2010.igem.org/Team:Imperial\_College\_London/Notebook](http://2010.igem.org/Team%3AImperial_College_London/Notebook)"

[![Powered by MediaWiki](/wiki/skins/common/images/poweredby_mediawiki_88x31.png)](https://www.mediawiki.org/)

[![Attribution 3.0 Unported](https://i.creativecommons.org/l/by/3.0/88x31.png)](https://creativecommons.org/licenses/by/3.0/)

* [Recent changes](/Special%3ARecentChanges "Recent changes")
* [What links here](/Special%3AWhatLinksHere/Team%3AImperial_College_London/Notebook "List of all wiki pages that link here [j]")
* [Related changes](/Special%3ARecentChangesLinked/Team%3AImperial_College_London/Notebook "Recent changes in pages linked from this page [k]")
* [Special pages](/Special%3ASpecialPages "List of all special pages [q]")
* [My preferences](/Special%3APreferences)

* [Printable version](/wiki/index.php?title=Team:Imperial_College_London/Notebook&printable=yes "Printable version of this page [p]")
* [Permanent link](/wiki/index.php?title=Team:Imperial_College_London/Notebook&oldid=382 "Permanent link to this revision of the page")
* [Privacy policy](/2010.igem.org%3APrivacy_policy "2010.igem.org:Privacy policy")
* [Disclaimers](/2010.igem.org%3AGeneral_disclaimer "2010.igem.org:General disclaimer")



[![](https://2010.igem.org/images/wiki.png)"](/Main_Page "Main Page")

* [Page](/Team%3AImperial_College_London/Results)
* [Discussion](/wiki/index.php?title=Talk:Team:Imperial_College_London/Results&action=edit&redlink=1)
* [View source](/wiki/index.php?title=Team:Imperial_College_London/Results&action=edit)
* [History](/wiki/index.php?title=Team:Imperial_College_London/Results&action=history)
* teams

* [Log in](/wiki/index.php?title=Special:UserLogin&returnto=Team:Imperial_College_London/Results "You are encouraged to log in; however, it is not mandatory [o]")

# Team:Imperial College London/Results

### From 2010.igem.org

[![](https://static.igem.org/mediawiki/2010/9/9f/ICparalogo.png)](https://2010.igem.org/Team%3AImperial_College_London)

[![](https://static.igem.org/mediawiki/2010/8/84/ICLlogo1.png)](http://www3.imperial.ac.uk/)

[![](https://static.igem.org/mediawiki/2010/7/7d/ICLlogo2.png)](https://2010.igem.org/Main_Page)

[![](https://static.igem.org/mediawiki/2010/7/7f/ICLlogo3.png)](http://www3.imperial.ac.uk/syntheticbiology)

[Department of Bioengineering](http://www3.imperial.ac.uk/bioengineering)

[Division of Molecular Biosciences](http://www3.imperial.ac.uk/molecularbiosciences)

* Project
* Plan
* Results
* Extras

Parasight  |  Parasite detection with a rapid response

* [Home](https://2010.igem.org/Team%3AImperial_College_London)
* [Tour](https://2010.igem.org/Team%3AImperial_College_London/Tour/Page_One)
* [Modules](https://2010.igem.org/Team%3AImperial_College_London/Modules)
* [Chassis](https://2010.igem.org/Team%3AImperial_College_London/Chassis)

+ [Human Practices](https://2010.igem.org/Team%3AImperial_College_London/Human_Practices)
+ [Schistosoma](https://2010.igem.org/Team%3AImperial_College_London/Schistosoma)
+ [Research](https://2010.igem.org/Team%3AImperial_College_London/Research)
+ [Diary](https://2010.igem.org/Team%3AImperial_College_London/Diary/Week_One)

* [Modelling](https://2010.igem.org/Team%3AImperial_College_London/Modelling)
* [Assembly](https://2010.igem.org/Team%3AImperial_College_London/Strategy)
* [Lab Diaries](https://2010.igem.org/Team%3AImperial_College_London/Lab_Diaries)

+ [Lab Protocols](https://2010.igem.org/Team%3AImperial_College_London/Protocol)
+ [Safety](https://2010.igem.org/Team%3AImperial_College_London/Safety)

* [Experimental Results](https://2010.igem.org/Team%3AImperial_College_London/Results)
* [Parts](https://2010.igem.org/Team%3AImperial_College_London/Parts/Favourites)
* [Achievements](https://2010.igem.org/Team%3AImperial_College_London/Achievements)

* [The Team](https://2010.igem.org/Team%3AImperial_College_London/The_Team)
* [Media](https://2010.igem.org/Team%3AImperial_College_London/Media/Videos)
* [School Workshops](https://2010.igem.org/Team%3AImperial_College_London/School_Workshops)
* [Software Tool](https://2010.igem.org/Team%3AImperial_College_London/Software_Tool)

+ [Acknowledgments](https://2010.igem.org/Team%3AImperial_College_London/Acknowledgments)
+ [Brainstorming](https://2010.igem.org/Team%3AImperial_College_London/Brainstorming)
+ [Glossary](https://2010.igem.org/Team%3AImperial_College_London/Glossary)
+ [Sitemap](https://2010.igem.org/Team%3AImperial_College_London/Sitemap)

|  |  |
| --- | --- |
| Experimental Results | [Exp 1](/Team%3AImperial_College_London/Results/Exp1 "Team:Imperial College London/Results/Exp1") |  [Exp 2](/Team%3AImperial_College_London/Results/Exp2 "Team:Imperial College London/Results/Exp2") |  [Exp 3](/Team%3AImperial_College_London/Results/Exp3 "Team:Imperial College London/Results/Exp3") |  [Exp 4](/Team%3AImperial_College_London/Results/Exp4 "Team:Imperial College London/Results/Exp4") |  [Exp 5](/Team%3AImperial_College_London/Results/Exp5 "Team:Imperial College London/Results/Exp5") |  [Exp 6](/Team%3AImperial_College_London/Results/Exp6 "Team:Imperial College London/Results/Exp6") |  [Exp 7](/Team%3AImperial_College_London/Results/Exp7 "Team:Imperial College London/Results/Exp7") |
| **Testing is a fundamental stage of the engineering design cycle and is a crucial part of charactrising BioBrick Standard Biological Parts so that other people can benefit from our work. We've compiled all our results on this page, detailing how the experiments were carried out and the significance of the data.** | |

|  |  |
| --- | --- |
| Experiment 1 | Optimum absorption wavelength | ![](https://static.igem.org/mediawiki/2010/e/e6/Spectra_of_Xyle_cells.jpg) |
| for catechol assays |
| **Aims of experiment |** In this experiment spectra from XylE expressing and XylE negative cells are compared after catechol addition. This allowed us to identify optimum absorption wavelength for quantification of output signal production. **Results |** Overlay of the two spectra from the cell cultures after catechol addition, reveals a broad peak appears with maximum at 380nm unique to XylE expressing cultures. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp1 "Team:Imperial College London/Results/Exp1") |

|  |  |
| --- | --- |
| Experiment 2 | The threshold value of catechol assay | ![](https://static.igem.org/mediawiki/2010/3/38/CS1.JPG) |
| **Aims of experiment |**  In this experiment we wanted to establish the threshold value at which a positive result (yellow color) would be detectable by the naked eye. **Results |**  The threshold concentration of catechol needed for a person to identify a yellow positive test was found to be 30-40μM. This result was fed back to the modelling team and allowed them to constrain the concentration values of catechol to be used. Furthermore, it facilitated the determination of conditions for modelling of 2-step vs 1-step amplification. Two step was shown to be better. The prediction for our system is that 2-step amplification output will be visible around 4 minutes earlier than the 1-step amplification system. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp2 "Team:Imperial College London/Results/Exp2") |

|  |  |
| --- | --- |
| Experiment 3 | Characterizing kinetic parameters of C(2,3)O in whole cells | ![](https://static.igem.org/mediawiki/2010/c/cb/IC_Assay_3_sept.jpg) |
| **Aims of experiment |**  In this experiment we attempt to characterize the kinetic parameters of catechol(2,3) dioxygenase enzyme, an existing registry part in whole cell cultures. **Results |**  Very useful information was acquired out of these assays of which the most important is the reaction profile of catechol(2,3)dioxygenase with catechol. The course of the reaction was analysed indirectly from the yellow product production over time at various initial catechol concentrations. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp3 "Team:Imperial College London/Results/Exp3") |

|  |  |
| --- | --- |
| Experiment 4 | Assaying cell-growth in presence of Catechol | ![](https://static.igem.org/mediawiki/2010/d/dd/XylE_M9_Growth_%28600%29.jpg) |
| **Aims of experiment |**  In order to assess possible effects of either Catechol or the breakdown product 2-hydroxymuconic semialdehyde, we performed growth assays under a variety of conditions. **Results |**  Experiments indicate that the breakdown products to have a strong negative effect on cell survival. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp4 "Team:Imperial College London/Results/Exp4") |

|  |  |
| --- | --- |
| Experiment 5 | Characterizing GFP-XylE gene product in whole cell cultures | ![](https://static.igem.org/mediawiki/2010/0/0d/GFPXylE.jpg) |
| **Aims of experiment |**  In this experiment we wanted to test if our custom designed biopart, GFP - XylE fusion protein, functions as expected. The specifications were set by the team to design an inactivated XylE reporter enzyme. **Results |**  Comparing reaction profile graphs of cell cultures of XylE expressing cells Vs GFP-XylE expressing cells showed that the designed reporter enzyme has more than 10 fold decreased activity in comparison to wild type reporter enzyme catechol(2,3)dioxygenase. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp5 "Team:Imperial College London/Results/Exp5") |

|  |  |
| --- | --- |
| Experiment 6 | In vitro characterization of C(2,3)O in cell lysate | ![](https://static.igem.org/mediawiki/2010/e/eb/Grapfit_curve_1.jpg) |
| **Aims of experiment |**  In this experiment we attempt to characterize in vitro the kinetic parameters of the enzyme catechol(2,3)dioxygenase (XylE) in cell lysate **Results |**  Construnction of Michaelis-Menten curve and determination of kinetic parameters of catechol(2,3)dioxygenase enzyme in E.coli. The calculated Km value - the substrate concentration at which velocity is half of the maximum, is 0.71mM of catechol. The Vmax is 3.37mM/min at x20fold dilution of cell lysate. |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp6 "Team:Imperial College London/Results/Exp6") |

|  |  |
| --- | --- |
| Experiment 7 | Characterization of Pveg promoter in r.p.u. | ![](https://static.igem.org/mediawiki/2010/f/ff/Untitled_5.jpg) |
| **Aims of experiment |**  Measuring the activity of BioBrick promoters using an in vivo reference standard. **Results |**  pVeg promoter in pSB1C3 vector, a high copy plasmid, has an 1.62 r.p.u value and in 3K3 vector, a low copy plasmid an 0.79 r.p.u. value. These values were derived by dividing signal from the production of HMS by the pVeg promoter population of cells by signal from the standard promoter J23101 (r.p.u value of 1). |
| [Learn More...](/Team%3AImperial_College_London/Results/Exp7 "Team:Imperial College London/Results/Exp7") |

|  |
| --- |
| Testing the Detection Module |
| Cloning the cell surface protein construct into *B. subtilis* has been problematic, which means we have not had time to upload testing results yet. However, we hope to have done this by the Jamboree. By increasing NaCl concentration to 1M, the electrostatic interactions will be disrupted and we can use a nickel column to bind the His-tagged surface protein thereby ensuring that it correctly attached to the cell wall. We then hope to test each cell surface linker by adding TEV protease and measuring how much AIP is cleaved. The linker that gives us the highest degree of cleavage will be used for the final surface protein for cleavage by the *Schistosoma* elastase. The characterisation of the linkers in the  [surface protein](/wiki/index.php?title=Team:Imperial_College_London/Modules/Detection_Module&action=edit&redlink=1 "Team:Imperial College London/Modules/Detection Module (page does not exist)") are definitely an area for future work, as the surface protein has a vast number of potential applications. For more information, check out our new  [Software Tool](/Team%3AImperial_College_London/Software_Tool "Team:Imperial College London/Software Tool")! |

|  |
| --- |
| Testing the Signaling Module |
| Due to problems with the synthesis of this construct, we only received it at the end of Week 16 and therefore have not had sufficient time to clone it and test it. However, this module would have been tested with a synthetic AIP, which has successfully been used in previous experiments, such as by [http://ukpmc.ac.uk/backend/ptpmcrender.cgi?accid=PMC40587&blobtype=pdf An unmodified heptadecapeptide pheromone induces competence for genetic transformation in *Streptococcus pneumoniae*] by Havarstein, Coomaraswamy & Morrison. This would have activated the signaling cascade, resulting in the phosphorylation of ComE, which would then be able to induce transcription of a reporter gene. However, this step would have been relatively straightforward and we have no reason to believe it would not function properly in *B. subtilis*, as like the organism from which the signaling system is taken (*S. pneumoniae*), it is Gram positive and has been shown to signal via AIPs in quorum sensing systems. |

Retrieved from "[http://2010.igem.org/Team:Imperial\_College\_London/Results](http://2010.igem.org/Team%3AImperial_College_London/Results)"

[![Powered by MediaWiki](/wiki/skins/common/images/poweredby_mediawiki_88x31.png)](https://www.mediawiki.org/)

[![Attribution 3.0 Unported](https://i.creativecommons.org/l/by/3.0/88x31.png)](https://creativecommons.org/licenses/by/3.0/)

* [Recent changes](/Special%3ARecentChanges "Recent changes")
* [What links here](/Special%3AWhatLinksHere/Team%3AImperial_College_London/Results "List of all wiki pages that link here [j]")
* [Related changes](/Special%3ARecentChangesLinked/Team%3AImperial_College_London/Results "Recent changes in pages linked from this page [k]")
* [Special pages](/Special%3ASpecialPages "List of all special pages [q]")
* [My preferences](/Special%3APreferences)

* [Printable version](/wiki/index.php?title=Team:Imperial_College_London/Results&printable=yes "Printable version of this page [p]")
* [Permanent link](/wiki/index.php?title=Team:Imperial_College_London/Results&oldid=205854 "Permanent link to this revision of the page")
* [Privacy policy](/2010.igem.org%3APrivacy_policy "2010.igem.org:Privacy policy")
* [Disclaimers](/2010.igem.org%3AGeneral_disclaimer "2010.igem.org:General disclaimer")