Team:Calgary/6 July 2010

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Revision as of 22:50, 6 July 2010 by Raidakhwaja (Talk | contribs)

Tuesday July 6, 2010

Happy Birthday Paul!!

Himika:

  • Today I reran the transformation that I did yesterday July 5th where I constructed E0040 and pSB1AC3 and plate it on chloramphenicol.
  • I also miniprepped the overnight cultures from last night. I10504 and I13507. The only cultures that grew were I10504 and I miniprepped 6 colonies from 1 plate.
  • I also constructed E1010 with B0015.
  • Procedure:
    • Insert
    • E1010 (pSB2k3) cut with EcoI/SpeI 5uL DNA
    • Buffer I was used 3.5uL
    • 0.5 uL of EcoRI and 0.5 uL of SpeI
    • 25.5 uL of water
    • Vector
    • B0015 (pSB1AK3) cut with EcoI/XbaI 5uL DNA
    • Buffer I was used 3.5uL
    • 0.5 uL of EcoRI and 0.5 uL of XbaI
    • 25.5 uL of water
  • The vector was phosphatased with 5uL water, 4uL Buffer and 1uL phosphatase. The solution was left in 37 degree Celcius for 30 minutes and heat killed for 10 minutes in 65 degree Celcius.
  • 5ul of the vector and the insert in a tube and add 10 ul of quick ligase buffer and 1 ul of quick ligase. The solution was left in room temperature for 10 minutes and transformed.
  • I also did research on the ibpA/ ibpB pathways in order to present on to Dr. Schryvers on Friday.
  • I also overnight cultured I10507 and plated the contstruct.

Raida: Today I worked on the Construction of R0040 (TetR promoter) infront of GFP construct to test the reporter. This construction is working as our positive control of verifying that our GFP reporter works.

  • "Restriction Enzyme Digest of Insert (R0040)"
    • 17.2 uL DNA
    • 3.5 uL React 1 Buffer
    • 0.5 uL EcoI, 0.5 uL SpeI
    • 13.5 uL of Water
  • "Restriction Enzyme Digest of Vector (I10507) Trial F"
    • 4.4 uL DNA
    • 3.5 uL React 2 Buffer
    • 0.5 uL EcoI, 0.5 uL XbaI
    • 26.1 uL of Water
  • "Restriction Enzyme Digest of Vector (I10507) Trial C"
    • 4.0 uL DNA
    • 3.5 uL React 2 Buffer
    • 0.5 uL EcoI, 0.5 uL XbaI
    • 26.5 uL of Water
  • "Restriction Enzyme Digest of Vector (I10507) Trial D"
    • 1.6 uL DNA
    • 3.5 uL React 2 Buffer
    • 0.5 uL EcoI, 0.5 uL XbaI
    • 28.9 uL of Water

At the end of the heat killing process, I added

    • 5 uL Antartic Phosphotase Buffer
    • 5 uL of Water and
    • 1 uL Antartic Phosphotase

It was placed in the water bath at 37 degrees Celcius for half an hour and then I ligated the insert and the vector using

    • 5 uL of Insert and 5 uL of Vector
    • 1 uL Quick Ligase
    • 10 uL 2x Quick Ligase Buffer

The construct is left in the fridge for overnight and transformation will be done tomorrow.

    • Today I also worked on the Ethics component of our Project. In particular I looked at the "feasibility" of Synthetic Biology to be manipulated for Bioterrorism Purpose.

No notebook page exists for this date. Sorry!