From 2007.igem.org

A Synchronized 2-gene Oscillator With Quorum Sensing

The aim of our team is to further understand the two-gene Oscillator, and couple this with the Repressilator system in order to understand time-varying conditions in the form of extrinsic driving from the environment and intrinsic rhythms generated within an organism itself. This includes specialized rhythm generators functioning in a coherent oscillatory state such as the cardiac pacemaker, also known as the sinoatrial node in mammalian hearts, and the circadian clock residing at the suprachiasmatic nuclei in mammalian brains.

This is a continuation of one of our projects presented last year. However, we intend to understand the effect of varying various conditions, such as controlling the cell densities, levels of AI (artificial inducer) by adding an AI analog, adding Tetracycline (DOX) to control LacI (Off-switch) and installing an AI inhibitor to control AI levels in the system.

1. Using Lac- MC4100 cells to obtain cleaner oscillations.
2. Changing cell densities and observing the effect on the oscillations.
3. Adding an AI Analog (AHL) into the system and observing the effect on the oscillations.
4. Adding Tetracycline (DOX), an inhibitor of the LuxR promoter, to the system and observing its effect.
5. Experimenting with cell divisions through an assay of optical density.
   
6. Synthesizing a "new I15004" brick to meet our needs.
7. Including an AI inhibitor (Aiia) in our 2-gene system to control the levels of AI produced.

J40001

Waiting to be synthesized. Almost 2 months late and still hasn't arrived from GeneArt...

I724000, the aiiA gene under the LuxR promoter

I724001, combination of I724000 and J40001, for ease of transformation (not yet made).

By coupling the Repressilator to the system, it is theorized that this more complex system can become highly stabilized if coupled with the 2-gene quorum-sensing oscillator, and once working side-by-side, the 2 systems will produce highly stable, smooth, sinusoidal oscillations.