Categories :

Home Team Sponsors Parts Submitted to Registry Image Gallery Leave a Message!

Project :

Overview Sensitivity Tuner --- Characterisation --- Modelling Colour Generators --- Carotenoids (Orange/Red) --- Melanin (Brown) --- Violacein (Purple/Green) The Future Safety

Notebook :

Week 1 Week 2 Week 3 Week 4 Week 5 Week 6 Week 7 Week 8 Week 9 Week 10

Team Logistics :

Protocols Stock List Research Shared Links and Help

A Reliable Amplification System

Introduction

The Cambridge 2007 iGEM team developed a Pops amplifier system using phage activators and promoters. The system works by using a Pops input to make an activator protein, as shown in the diagram from their wiki below, which then binds to a promoter and generates a Pops output.

In order to quantify the ratio between Pops in and Pops out, the team built the following construction on the high copy plasmid pSB1A2, with mRFP and GFP as Pops reporters and 15 total combinations of different activators and reporters.

They successfully quantified the Pops amplification factors for each activator/promoter combination after arabinose induction. However, after data analysis they pointed out two phenomena that required further investigation:

1. Cultures transformed with the amplifier constructs showed reduced growth which increased in severity with increased arabinose induction. It was suspected that the activators were toxic to the cells at high concentrations.

2. RFP was an unreliable reporter. For some reason, RFP appears to be more stable when its gene is downstream of another gene in a polycistronic transcript compared to when its gene is directly downstream of a promoter.

The Cambridge 2009 iGEM team hopes to debug this system by concentrating on these two problems.

Project

Starting off with 2007

We began by recreating the 2007 team's data to see if we would encounter the same problems.

The Experiments

Results