Difference between revisions of "Team:Exeter/InterLab"

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600nm absorbance of all samples was recorded as follows:
 
600nm absorbance of all samples was recorded as follows:
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Interlab data (ludox part)  
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<b>Interlab data (ludox part)</b>
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The final calculated results were:
 
The final calculated results were:
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Interlab data (ludox part from excel sheet with calculations)  
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<br>
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<b>Interlab data (ludox part from excel sheet with calculations)<b>
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The corrected Abs600 was calculated by subtracting the H20 reading. The reference OD600 was defined as measured by the reference spectrophotometer (provided by iGEM HQ). The correction factor to convert measured Abs600 to OD600 was hence the Reference OD600 divided by Abs600. In order to maintain consistency of the results, we used the same plates and volumes for the subsequent steps.  
 
The corrected Abs600 was calculated by subtracting the H20 reading. The reference OD600 was defined as measured by the reference spectrophotometer (provided by iGEM HQ). The correction factor to convert measured Abs600 to OD600 was hence the Reference OD600 divided by Abs600. In order to maintain consistency of the results, we used the same plates and volumes for the subsequent steps.  
 
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Revision as of 14:27, 30 October 2017

In 2017, the iGEM InterLab Study asked teams to help developing a standardised curve of fluorescence measurements and to measure the relative strength of some unknown devices with different RBSs:

  • Device 1: BBa_J364000
  • Device 2: BBa_J364001
  • Device 3: BBa_J364002
  • Device 4: BBa_J364003
  • Device 5: BBa_J364004
  • Device 6: BBa_J364005
  • Positive Control: BBa_ I20270
  • Negative Control: BBa_R0040

This involved measuring the fluorescence/OD600 of cultures of E. coli DH5α transformed with each construct to compare the relative expression per cell. By asking all iGEM teams to use the same calibration process for the instruments used for the measurements and comparing the results to a measured standard curve, the relative units of fluorescence can be converted to absolute units. We started our contribution by resuspending the appropriate DNA from the iGEM distribution kit.

OD600 reference point

The experiment was carried out on the 10th July 2017. The first step was to use LUDOX-S40 as a single point reference to obtain a ratiometric conversion factor to transform the absorbance data into a standard OD600 measurement. In order to do this, we used a 96-well plate. 100μl of LUDOX were added into wells A1, B1, C1 and D1 of the 96-well plate. The same amount of H2O was added into wells A2, B2, C2 and D2. The parameters used for the TECAN plate reader were:

  • Excitation wavelength: 485nm
  • Emission wavelength: 530/30nm
  • Gain: 45

600nm absorbance of all samples was recorded as follows:
Interlab data (ludox part)
The final calculated results were:
Interlab data (ludox part from excel sheet with calculations)
The corrected Abs600 was calculated by subtracting the H20 reading. The reference OD600 was defined as measured by the reference spectrophotometer (provided by iGEM HQ). The correction factor to convert measured Abs600 to OD600 was hence the Reference OD600 divided by Abs600. In order to maintain consistency of the results, we used the same plates and volumes for the subsequent steps.

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