Purdue/2 September 2008

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Restriction Digest of SOS and LacZ

1. Mixed digestion solution:

  • 50uL DNA
  • 7uL NEB2
  • 10.5uL ddH2O
  • 0.5uL BSA
  • 1uL of each enzyme
    • SOS: EcoRI + SpeI
    • LacZ: EcoRI + XbaI
    • This makes SOS the small part cut out (192 bp) and put into the LacZ plasmid (~5000 bp...=LacZ + most of plasmid)

2. Put mixtures at 37C for 1 hour, 25 min.

3. Run through Agarose gel w/ DNA ladder, DNA loading dye (Bio-RAD)

  • Mixed 7uL of loading dye w/ each 70uL tube
  • 10uL ladder in lane 1
  • Lanes 2-4: 20uL LacZ in each lane
  • Lanes 5-7: SOS
  • 120V, 45 min.

4. Overnight stain w/ Fast Blast (5x)


Edited by Janie Stine

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