User:Karmella Haynes/Notebook/BioBrick cloning/2015/04/28

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04/28/15

  • Ryan - minirpeps & digests
  • Line item 2


Minipreps

  • Use Sigma GenElute; elute with 75 μL elution solution
  • BraR/MRV colonies - picked 4 additional colonies from original cloning plate
  • Cut w/ E/X
  1. Bra/MRV 3
  2. Bra/MRV 4
  3. Bra/MRV 5
  4. Bra/MRV 6
  • promoter/Regulator/MRV colonies
  • Cut w/ KpnI/EcoRI (BbsI drop-in of promoter destroys KpnI site); empty vector will produce 2 bands, successful clones will be linearized (1 cut)
  1. pAub/AubR/MRV
  2. pBja/BjaR/MRV
  3. pRpa/RpaR/MRV


Reagent Volume Expected:
1-4. Bra/MRV = ###
Empty MRV E/X = ###
5. pAub/AubR/MRV = ###
6. pBja/BjaR/MRV = ###
7. pRpa/RpaR/MRV = ###
Empty vector = ~###, 971
DNA(plasmid) 3.0 μL
10X buffer 1.5
enzyme 1 1.0
enzyme 2 1.0
dH2O 8.5
  15 μL --> 37°C/ ~15 min.



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