iGEM2005 project

Aims & Objectives: The ultimate aim of the project is to display a significant degree of control over not only the individual cells motion but also the population as a whole in an attempt to demonstrate spatiotemporal pattern formation. Ideally, we'd like to demonstrate and design a morphogenic system

Biological Analogue

The main biological analogue would be: Transient gene expression leading to permanent activation of behaviour

Inspiration

We have taken inspiration from:

• Weiss' pulse generating network
• Alice & Chris' pretty cool flipping/recombinase switch
• The controllable chemotaxis vision of James & Russell.
• What it does, how it works

Chemotaxis is (pretty much) irreversibly switched on in a strain not normally displaying this behaviour

• This bacterium is a MalE knockout strain, procured from Sweden/Japan/somewhere exotic. James & Russell plan to rescue Chemotaxis using the MalE gene (encoding the Maltose Binding Protein, MBP) or a rapidly degrading version (MalE 31).
• Alice & Chris have been working on using a site-specific recombinase as a genetic switch by trapping a promoter between two recombination sites and using their 'controllable' Hin recombinase to play with its orientation. This way they can generate mutually exclusive expression of two gene sets.
• Russell & James have also gotten their grubby mits on the pMalT promoter, which is activated by the MBP, when bound to maltose. By using this to control a reporter gene, they think they can show when a bacterium can 'see' maltose.
• Eva and the other James have been faffing about with the pulse generation idea and if they can incorporate a nice way to turn on the recombinase for a short amount of time - hopefully we can get the majority of the E. coli to display chemotaxis at once. Obviously there is a risk of the recombinase getting over-zealous and flipping the promoter back.

Using an external stimulus (yet to be decided), we will stimulate a pulse of recombinase expression - hopefully just enough for our promoter to flip orientation once, leading to the expression of the Maltose binding protein in a MalE knockout strain - .: exhibiting chemotaxis. The bacterial traffic lights get another look in here too, with mCherry expression being the default state, GFP expression replacing this after recombination and mOrange expression under a MalT promoter.

I'm pretty excited about this - when we add our stimulus we're going to get some of the bacteria starting to move and change colour, and those that don't will stay still and red. We can view the dynamics at macro and micro scales, even bacteria that flip their promoter too often won't be too annoying, they'll just be slower movers.