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From 2010.igem.org

Contents 1 July 5th 2 July 6th 3 July 7th 4 July 8th 5 July 9th

July 5th

• The university was closed due to the holiday on July 4th.

July 6th

• Back to work!

• Tomorrow:

  - Retransform:
       o All lux promoters from kit plates.
  

• Grow for tomorrow from stock:

       - R0061
       - J24679
       - R1062
       - K112022
       - K091107
       - K091146
       - K145150

• Cleaned cuvettes for transformations.

• Worked on Human Practices survey.

July 7th

• Flasks minipreped:

      - J24679
      - R1062
      - K112022
      - K091107
      - K091146
      - K145150

• Sent to sequencing:

 1) P0451
 2) K112022
 3) J24679
 4) K091146
 5) K145150 
 6) R0010
 7) R1062
 8) K091107

• Digests:

  - J24679 (ES+phosphatase)
  - P0451  (XP+phosphatase)
  - K274100(ES+phosphatase)
  - S01414 (ES+phosphatase)
  - K274100(XP+phosphatase)
  - B1002  (EX)
  - B1006  (EX)
  - R0010  (SP)
  - B1002  (EX)
  - B1006  (EX)
  - J23100 (SP)

• Gel order:

  - Ladder
  - B1006
  - J23100
  - R0010

All three bands appeared around the base pairs that we wanted then at so the 3 bands were cut out and then gel purified.

• Ordered 3 primers.

• Ligations:

  Insert          Vector

1) J24679 B1006 2) P0451 R0010 3) K274100 B1006 4) S01414 B1006 5) K274100 J23100

July 8th

• Colony PCR:

 1) P0451+R0010 A-D
 2) K274100+B1006 A-D 
 3) K274100+J23100 A-D
 4) J24679+B1006 A-D

• Digest:

 - J24679 (ES+phosphatase)
 - K274100(ES)
 - R0010 (SP)
 - B1002 (EX)
 - K112022 (XP)
 - K112808 (XP)

• Gel order:

 Upper-digests
    - Ladder
    - K274100
    - K112808
    - K1120222
    - B1002
 Lower-PCR
    - Ladder
    - 1 A-F
    - 4 A-E

• We forgot to put the Taq polymerase and so as you can see, nothing showed up in the gel.

July 9th

• To grow:

  -C0261

• Transform:

  -C0261 from kit plate 2006 
  -R0051

• Gel order:

  -Ladder
  -1A-D (K274100+B1006)
  -2A-D (P0451+R0010)

  -Ladder
  -3A-3D (K274100+J23100)
  -4A-D  (J24679+B1006)

- 2B, 2C, 4A, and 4C were grown while 2A was digested. 
- However not a single band was found above 500 base pairs for the other 3 that were colony PCR-ed

• Sent to sequencing:

  1)J24679+B1106 (E)
  2)P0451+R0010 (A)
  3)P0451+R0010 (F)

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