IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/09/21

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dspB Track

Miniprep O/N cultures C,D,G,H

  • Protocol: Rafael's Alkaline Lysis Protocol
  • Resuspend DNA in 70uL of water
  • Nanodrop:
    • [C] = 1777.0ng/uL
    • [D] = 1318.5ng/uL
    • [G] = 1373.8ng/uL
    • [H] = 842.5ng/uL

Restriction Digest

  • Protocol: "digest" protocol from biobrick
Restriction Digest
REAGENTS1 RXN VOLUME (uL)
Buffer 22
BSA0.2
DNA10
ddH207.8
Total20
  • x = amount of DNA
  • DNA:
    • Cut terminator (suffix) at E/X
    • Cut D and G at E/S
  • Add 1uL of each appropriate enzyme

Digest for 2 hours; start at 3:20pm

Gel verification colony PCR products

  • Protocol: gel verification protocol in Protocol (SOP)
  • Machine conditions: 0.5x TBE buffer, 100V, 60min

Gel orientation:

Gel orientation
DG100bp ladderE/X term1 kb ladder

Results:

  • Digestion failed.

Re-Restriction Digest

  • Protocol: "digest" protocol from biobrick
Restriction Digest
REAGENTS1 RXN VOLUME (uL)
Buffer 22
BSA0.2
DNA10
ddH207.8
Total20
  • DNA:
    • Cut terminator (suffix) at E/X
    • Cut D and G at E/S
  • Add 1uL of each appropriate enzyme

O/N digestion starting at 9:30pm



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