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Team:Cambridge/Project/Amplification
From 2009.igem.org
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| - | We began by recreating the 2007 team's data to see if we would encounter the same problems. We used the E. coli host strain BW27783. This host strain constitutively expresses arabinose transporters and is unable to metabolize arabinose, making it an ideal host for arabinose titration experiments. | + | We began by recreating the 2007 team's data to see if we would encounter the same problems. We used the E. coli host strain BW27783. This host strain constitutively expresses arabinose transporters and is unable to metabolize arabinose, making it an ideal host for arabinose titration experiments. We have the advantage over the 2007 team in that we have a better plate reader that is able to take OD600 absorbance readings at the same time as taking RFP and GFP absorbance readings. |
== Experiments == | == Experiments == | ||
Revision as of 09:32, 6 August 2009
Categories :
Project :
-
Overview
Sensitivity Tuner
--- Characterisation
--- Modelling
Colour Generators
--- Carotenoids (Orange/Red)
--- Melanin (Brown)
--- Violacein (Purple/Green)
The Future
Safety
Notebook :
Team Logistics :
A Reliable Amplification System
Introduction
Cambridge iGEM 2007
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.
Further work - Cambridge iGEM 2009
However, after data analysis they pointed out one 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.
The Cambridge 2009 iGEM team hopes to debug this system by concentrating on these two problems.
Recreating Previous Work
We began by recreating the 2007 team's data to see if we would encounter the same problems. We used the E. coli host strain BW27783. This host strain constitutively expresses arabinose transporters and is unable to metabolize arabinose, making it an ideal host for arabinose titration experiments. We have the advantage over the 2007 team in that we have a better plate reader that is able to take OD600 absorbance readings at the same time as taking RFP and GFP absorbance readings.
