IGEM:IMPERIAL/2007/Projects

Vincent 17:18, 9 July 2007 (EDT): A page to describe briefly the projects the team is exploring, and to link to their documentation.

Project Description

Engineering biology 'in-veso' - Imperial College iGEM 2007 team

The aim of our project is to investigate the use of cell-free chassis to realise new potentials for simple constructs. To date, work on synthetic biology has been done using the chassis of bacterial cells. However the use of living, replicating engineered bacteria poses a huge limitation for applications in the medicine and food industries for reasons of public safety. To solve this problem, our team decided to introduce cell-free expression systems as a new chassis to the field of synthetic biology. For comparison, we are going to use the same DNA constructs in three different chassis. The first chassis is the traditional E. coli cells. Second, we will use in-vitro systems composed of E. coli cytoplasmic extracts. Third, the E. coli cytoplasmic extracts will be encapsulated in phospholipid vesicles to form in-veso systems. Using vesicles to compartmentalize the transcription-translation of plasmid genes can potentially prolong the lifespan of the systems by allowing exchange of nutrients and waste materials between the expression machineries and the surrounding feeding solution. To illustrate the feasibility of cell-free chassis, we aim to assemble genetic circuits in-veso for two exciting applications. One of the applications is Cell By Date, a device that determines when food is spoilt more accurately than estimates used to construct the printed sell by date. It exploits the thermal dependence of the rate of expression of a simple reporter system. The other application is Infector Detector, a system that detects biofilms on catheters and produces a fluorescent output. Biofilms are a serious problem in hospitals as they cause infections and are resistant to antibiotics. Our Infector Detector makes use of the Lux quorum sensing system to detect AHL secreted by the bacteria constituting a biofilm. This is analogous to eavesdropping on the communication between biofilm-forming bacteria. Engineering biological systems in veso eliminates the need for bacterial contact with either food or medical devices, this demonstrates the advantages of our cell-free chassis over the traditional E. coli cells.

DNA Consructs

DNA Constructs

DNA constructs for all our projects.

Lab Notebook

Follow the link above to the team's lab notebook

General Protocols

Protocols being used by all projects. Examples:

• Plasmid preparation
• Plasmid extraction
• Restriction digest
• Ligation

Reporters

Any reporters that might be used for the projects

Equipments needed

1. LB media
2. LB+ Amplicillin plate x 30
3. LB+ Kanamycin plate x 30
4. LB+ Amp+ Kn x 15
5. 37oC shaking incubator x 1
6. 4oC freezer x 1
7. Table top centrifuge x 3
8. 4oC table top centrifuge/ cold room/ fridge
9. UV Spectrometer x 2
10. Gilson pipettes (1000, 200, 20, 10, 2) x5 each
11. Tips (yellow and blue) x 20 boxes each
12. Small tips x 5 boxes
13. Eppendorf tubes
14. Centrifuge tubes
15. Machine to check for purity of DNA (260/280)
16. 37oC water bath
17. Metal block incubator
18. Innoculation loops x 10 packets
19. 15oC water bath
20. Agarose gel equipments x 3
21. TBE x 1 litre of 50x TBE
22. Microwave oven
23. Camera
24. Blades (for cutting agarose gel) x 3
25. Band detection machine
26. EB buffer (Tris-Cl + EDTA)
27. Gloves
28. Whiteboard markers
29. Paper towels
30. 10ml pipettes x 50
31. Pipette fillers x 5