User:Karmella Haynes/Notebook/BioBrick cloning/2010/05/28

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05/28/10

  • ✓ Minipreps: MV1 (2), MV7 (2) (no methylation)



Minipreps
> Check with BsaBI/ BstBI digests (methylation sensitive)
> Use old preps from DH5α as a control

Reagent Volume Expected:
1,2. MV1 (dam-) = 823, 4605
3. MV1 (dam+) = uncut
4,5. MV7 (dam-) = 823, 3919
6. MV7 (dam+) = uncut
Hover name
15 μL/lane; 1% agarose
DNA(plasmid) 2.0 μL
10X buffer 1.5
EcoRI 1.0
PstI 1.0
dH2O 9.5
  15 μL

--> 60°C (BsaBI)/ 1 hr.; 65°C (BstBI)/ 1 hr.
--> Success! C2925 cells produce non-methylated plasmids. Proceed w/ cloning hygro (PCR) into these plasmids.




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