IGEM:IMPERIAL/2007/Projects/Hrp System/Testing

=Hrp Characterisation: Testing=

 Introduction Specifications Parts Devices Testing Application Notes  References </ul> <br style="clear:both">

Proposed Schedule

Motivation

 * The motvation behind the characterisations carried out is to create well characterised parts and devices, and to characterise in terms of; GFP molecules synthesized cfu -1 sec-1.

Dependencies
General tasks to be completed for each of our constructs:
 * 1) Amplifying the biobricks
 * 2) Cloning of the constructs
 * 3) Transformation
 * 4) Growing Cultures
 * 5) Fluorometer
 * 6) Measuring Inputs - GFP downstream of Promoters
 * 7) Measuring Outputs- GFP downstream of Devices
 * 8) Calibration

2. Cloning of the Constructs
Link to General Protocol

3. Transformation
Link to General Protocol

5. Fluorometer

 * The fluorometer used in these experiments is 'The Twinkle'.
 * This machine is a plate reader fluorometer that can measure 96 well plates.
 * Using this machine it is hoped that absorbance at 600nm and the fluorescence emission of acGFP can be measured at set time intervals.
 * The machine is being installed 3rd of August 10.30am.


 * The general principle is to measure the fluorescence into the device and out of the device, this allows characteristics such as transfer function to be calculated.
 * To enable us to measure inputs and output, two separate experiments will be carried out, one for the inputs fluorescence and one for the output of fluorescence.


 * A  Hrp Protocol can be found here. This is a general protocol and can be applied to the various devices and parts to be characterised.
 * Two specific fluorescence measurements will be made for each device:
 * 1) Input -The input of fluorescence will be measured at varying inducer concentrations, to give the value of GFP molecules synthesized cfu -1 sec -1 into our device at specific inducer concentrations.
 * 2) Output -The output of fluorescence will be measured at varying inducer concentrations and at various time intervals. The inducer concentration will have a known GFP molecules synthesized cfu -1 sec -1 input and so we will be measuring the time response and level of output of our device.

6. Calibration

 * The two measurements that will be made for the various devices are fluorescence emission and absorbance.
 * The aim of the characterisation is to use these two measurements in order to create a more generic unit to allow for modular design and easier repetition of experiments. The unit that has been chosen to be used is GFP molecules synthesised cfu -1 sec-1.
 * In order to convert our raw data into these units, two sets of calibration curves are needed.
 * 1) Absorbance
 * 2) Fluorescence
 * Here is a link to an explanation of the Calibration Curves