IGEM:MIT/2006/Notebook/2007-7-26

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Things to Do

1. Miniprep J45120, J45181, J45995, and J45996- DONE

2. Sequence J45120, J45181, J45996, and J45996 before 10 AM- DONE

3. Set up plate reader experiment using the following protocol- DONE:

  • Add 1 mL overnight cultures to 24 mL LB (plus antibiotics) media in 125 mL flask
  • Put on shaker at 220 RPM for 50 mins
  • Add 200 uL of culture plus LB AMP/KAN to three wells each in triplicate

C1-C3 = J45995

C4-C6 = J45996

C7-C9 = R0040.E0840

C10-C12 = B0015

D1-D3 = LB AMP/KAN

4. Talk with Drew about abstract

5. Talk with Tom about media- DONE (wants to meet tomorrow after doing research)

6. Check Dr. Knight's calculations on media ingredients- DONE (nothing wrong besides roughly a gram difference of K2HPO4)

7. Do some research on media yourself- DONE (email Tom this paper http://jb.asm.org/cgi/reprint/89/4/996?view=long&pmid=14276127)

8. Miniprep stock LC of J45120 since the original miniprep gave a low yield of DNA (10.1 ng/uL)- DONE

9. Submit J45120 for sequencing- DONE

10. Add methyl salicylate to 25-mL cultures left over from plate reader experiment (0 ppm, 0.5 ppm, 1.5 ppm, 2.5 ppm)- DONE

Results from smell-a-rama

Eleven people surveyed.

  • 0/11 smelled mint in the 0 ppm culture
  • 5/11 smelled mint in the 0.5 ppm culture
  • 6/11 smelled mint in the 1.5 ppm culture
  • 10/11 smelled mint in the 2.5 ppm culture

So, it seems valid to say that, based on a small sample size, the threshold for smelling methyl salicylate seems to be between 1.5 ppm and 2.5 ppm or between 11.6 uM and 19.3 uM. This value is a small fraction of the 2 mM of salicylic acid added as a precursor to the product device.

1.5 ppm = 1.5 uL methyl salicylate/ 1 L solution

1.5 uL * 1.174 g / 1000 uL = .001761 g / 1 L solution

.001761 g / 152.1494 g/mol = 11.6 uM

2.5 ppm = 19.3 uM

Isoamyl acetate is said to have an odor threshold of 0.22 ppm, or 1.48 uM, http://www.blackwell-synergy.com/doi/abs/10.1111/j.1600-0668.1998.t01-3-00004.x

0.22 ppm = .22 uL isoamyl acetate / 1 L solution

.22 uL * 0.876 g/ 1000 uL = .00019272 g

.00019272 g / 130.19 g/mol = 1.48 uM

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