Team:Harvard/Dailybook/Week3/Widgetry

From 2008.igem.org

Contents

Goals for this week

  1. Make Orianna's growth media
  2. construct more chambers to run parallel tests
  3. set up Keithley for multiple tests
  4. grow mtrA, mtrB,wt, E. coli in liquid cultures
  5. negative control E. coli in LB/lactate
  6. standardize tests (OD, bubbling, etc)

Monday: June 7, 2008

  • E. Coli Negative Control in LB

EColi LB.JPG

EColi LB O2.JPG

Tuesday: June 8, 2008

  • Constructed two new MFCs
  • grew mtrA, mtrB, wt, and E. coli in LB+lactate liquid cultures
  • Ran negative control with E. coli in NaPIPES media


EColi NaPIPES.JPG

Wednesday: June 9, 2008

  • Made Orianna's growth media
First followed recipe exactly: pH = 3.4
  • Added NaOH to neutralize media
Missing amino acids
  • Grew wt Shewie, mtrA, mtrB, and mtrC/omcA in Orianna's growth media
    • overnight cultures
    • RESULTS:
OD readings:
wt: -0.001
mtrA: 0.001
mtrB: 0.003
mtrC/omcA: 0.002
CONCLUSION:
Orianna's growth media isn't working. Go see Colleen for possible sources of error.
  • OD readings from liquid cultures grown in LB + lactate growth media
    • wt: 2.489
    • mtrB: 1.711
  • Ran MFC overnight with mtrB and wt Shewie strains.
    • added strains first without lactate
    • waited until chambers became anaerobic (color-change) to add lactate
    • wt cell chamber denser than mtrB cell chamber
    • N2 stopped in wt chamber. turned off N2 in mtrB chamber to mirror this change

Mfc wt mtrB.png

  • Experiment continued
    • replenished lactate in wt chamber
    • fluctuations may have resulted from air/N2 flow

Mfc wt mtrB cont.png

Thursday: June 10, 2008

  • Continued running experiment from Wednesday
  • Purchased pipe cleaners, resistors
  • Made OGM with potassium PIPES

Friday: June 11, 2008

Lab

  • set up three chamber system
  • tested wt, mtrA, negative control:
    • resistances:
      • wt:
      • mtrA:
      • none:

Mfc overnight1.png

465 s: injected Shewie

3500 s: injected Lactate & stopped N2

Mfc overnight2.png

  • Nitrogen stopped bubbling somewhere between 70,000 & 80,000


Mfc overnight3.png

Conclusion

  • Day 1, current production of wt and mtrA Shewie match that in literature
    • When wt peaks at around 275 microamps, mtrA Shewie is producing 25-50 microamps
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