Imperial College/Courses/Spring2008/Synthetic Biology/Part Measurement and Characterisation/Day 5

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Spring 2008 - Introduction to Synthetic Biology


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Wet Lab: Part Measurements and Characterisation


1. Take out 1ml from each of the flasks left overnight in the 37°C shaking incubator from Thursday with a p1000 Gilson and transfer to 1ml cuvettes. Label the cuvettes “995” and “996”. One of the pair should measure the OD600 of both samples and record the readings. Before measuring the OD, dilute the sample in LB medium! Label this sample with 24h 995 and 24h 996 dependent on when the experiments was started the day before. The other of the pair should also take out 1ml from each flask but transfer the aliquots to Eppendorf tubes labelled “24h 995” and “24h 996”. Also, label the tubes with your initials. Leave these tubes in your ice bucket as these will be used for fluorescent measurements later on in the day.

GFP standard curve Serial dilutions of purified GFP protein (27.8kDa) (0.6mg/ml ~ 20µM) in PBS

  • 0 nM: 0 µl GFP + 200 µl PBS
  • 10nM: 0.1 µl GFP + 199.9 µl PBS
  • OR dilute the GFP 1:10 (1µl GFP + 9µl PBS) and use
    • 1µl GFP 1:10 + 199 µl PBS
    • 100 nM: 1µl GFP + 199 µl PBS
    • 300 nM: 3 µl GFP + 197 µl PBS
    • 600 nM: 6 µl GFP + 194 µl PBS
    • 900 nM: 9µl GFP + 191 µl PBS
    • 1200 nM: 12 µl GFP + 188 µl PBS
    • 1500 nM: 15 µl GFP + 185 µl PBS
    • 2000 nM: 20 µl GFP + 180 µl PBS