Team:Harvard/Dailybook/Week8/Widgetry

From 2008.igem.org

Contents

Goals for this week

  1. Run co-culture test with DH5α, DH5α/pUC19, and Shewie
  2. Move chambers to the "dark" to prevent a reaction from happening with light and oxygen
  3. Test light-repressible system on plates


Monday: August 11, 2008

  • cleaned chambers
  • moved setup to a cabinet to have system in the "dark"

DH5α, DH5α/pUC19, and Shewie co-culture experiment setup

  • Chamber 1: DH5α + wt Shewie + Lactose
  • Chamber 2: DH5α + wt Shewie + Lactate (pos. control)
  • Chamber 3: DH5α + wt Shewie (neg. control)
  • Chamber 4: DH5α/pUC19 + wt Shewie + Lactose
  • Chamber 5: DH5α/pUC19 + wt Shewie + Lactate (pos. control)
  • Chamber 6: DH5α/pUC19 + wt Shewie (neg. control)
  • Chamber 7: wt Shewie + Lactose (neg. control)
  • Chamber 8: DH5α + Lactose (neg. control)
  • Chamber 9: DH5α/pUC19 + Lactose (neg. control)


Tuesday: August 12, 2008

DH5(alpha) & DH5(alpha)/puC19 + wt Shewie Co-Culture Results

First Half

Dh5a first.jpg

  • Notes:
    • At t = 5000 s, all cells added
    • At t = 90000 s, all injections done
    • At t = 155000 s, Lactose injections on Chambers 1 and 4

Second Half

Dh5a second.jpg

  • Notes:
    • Computer restarted in middle of night, t=0 in second graph is 7 hours after end of first graph
    • At t = 925000 s,
Lactose injections on Chambers 2 and 5
Lactate injections on Chambers 3 and 6

Concerns

  • Repeatable current production in DH5(alpha) alone
    • Possible contamination?
  • Distinguishing features to tell current production of DH5(alpha) and DH5(alpha)/pUC19 apart
    • After first lactose injection, the current production of the two have noticeable difference in slope
    • However, initially after second lactose injection, the graphs overlap

Wednesday: August 13, 2008

RU1012 Light Repressible system experiment

  • Streaked plates.
  • All Cm/Amp plates with Xgal
Dark
  • Plate 1: RU1012 w/ plasmids
  • Plate 2: RU1012
  • Plate 3: Negative control (No bugs)
Light
  • Plate 1: RU1012 w/ plasmids
  • Plate 2: RU1012
  • Plate 3: Negative control (No bugs)


Results

RU1012 with Plasmid
RU1012 plasmid.JPG
  • Notes:
    • Plate on left was grown under light
    • Plate on right was grown in the dark
    • Possible repression seen in bottom left corner of the plate, but seems leaky
RU1012
RU1012 light.JPG RU1012 dark.JPG
  • Notes:
    • Plate on left was grown under light
    • Plate on right was grown in the dark
    • Exhibited same behavior as RU1012 with plasmid
Negative Control
  • Nothing on plates for both light and dark
Conclusions
  • Hard to distinguish if it worked with streaked colonies
  • May work better with lawns

Co-Culture Experiment Follow-up

  • When disassembling the fuel cells, took the chamber media and spread them on Xgal plates
    • Way of determining if contamination occured
Controls are not expected to turn blue

Thursday: August 14, 2008

RU1012 Light Repressible system experiment

  • Spread plates to try to create lawn.
  • All Cm/Amp plates with Xgal
Dark
  • Plate 1: RU1012 w/ plasmids
  • Plate 2: RU1012
  • Plate 3: Negative control (No bugs)
Light
  • Plate 1: RU1012 w/ plasmids
  • Plate 2: RU1012
  • Plate 3: Negative control (No bugs)

Results

RU1012 with plasmid under light
RU1012 plasmid light.JPG
  • Note:
    • Colonies had specks of blue, but not very noticeable
RU1012 with plasmid in the dark
RU1012 plasmid dark.JPG
Conclusion
  • Possibly light is too strong? Explaining no growth in middle of plate grow under light

Friday: August 15, 2008

RU1012 Light Repressible system experiment

  • Repeated previous day's experiment exactly
  • Placed light further away

Co-culture Experiment Follow-up

  • All plates turned blue
  • Control chambers very likely were contaminated
  • Will explore possibilities of autoclaving chambers