IGEM:IMPERIAL/2007/new pages/Data anaylsis
Conversion of Data
Using our calibration cuvre of GFPmut3b in our in vitro chassis we are able to convert the fluorescence units into a number of different units. We have looked at how we can convert between four different units:
- Fluorescence to Concentration of GFPmut3b
- Fluorescence to Moles of GFPmut3b
- Fluorescence to Molecules of GFPmut3b
- Fluorescence to Molecules of GFPmut3b per plasmid
There is a massive advantage for being able to interconvert between units. As of yet there is no standardised units for in vitro, for in vivo work on the F2620 used the standard units molecules of GFPmut3b synthesised per second per cfu (colony forming units). The units above allow a variety of different units to be used including a normalised unit based upon the number of DNA plasmids present in the chassis.
There are certain constraints to using our calibration curve:
- First the total volume must be kept constant as volume can affect fluorescence, we chose a volume of 60µl.
- The type of cell extract may also be important, this is for Promega S30 Cell Extract
- The DNA concentration used for the calibration curve could affect the fluorescence measure, therefore we standardised the DNA used in our experiments to 4µg.
- Finally because of fluorometer variation then at least the same conditions must be used if not the same fluorometer.
The diagram below shows the various conversions that can be carried out: