User:Karmella Haynes/Notebook/BioBrick cloning/2015/05/29

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05/29/15

  • David Tze - minpreps & digests
  • Ben - sequencing order
  • Cas-tone - sequencing order


David Tze - minpreps & digests

  • Minipreps
    • Sigma GenElute kit, elute w/ 75 μL elution sln.
  • Digests
    • Check with E/S digests; cannot use PstI because PLrigid & PLrigid4 contain PstI sites
Reagent Volume Expected:
1. DT001 = 2668, 252
Empty PLflex = 2668, 95
2. DT002 = 2656, 432
Empty PLflex4 = 2656, 274
3. DT003 = size
Empty PLrigid4 = #
4. DT004 = size
Empty PLrigid4 = # 		Hover name
15 μL/lane; 1% agarose; Ladder
DNA(plasmid) 2.0 μL
10X buffer 1.5
EcoRI 1.0
SpeI 1.0
dH2O 9.5
  15 μL --> 37°C/ ~15 min.

CONCLUSIONS:

  1. DT001 - success. Keep clone #2 (higher concentration). Sequence-verify.
  2. DT002 -
  3. DT003 -
  4. DT004 -

Sequencing order

  • 1-4 - Ben
  • 5-16 - Cas-tone
  1. DBN007-1 - VF2
  2. DBN007-1 - VR
  3. DBN007-2 - VF2
  4. DBN007-2 - VR
  5. H3140-1 - CMVf
  6. H3140-1 - DD123
  7. H3140-2 - CMVf
  8. H3140-2 - DD123
  9. H3160-1 - CMVf
  10. H3160-1 - DD123
  11. H3160-2 - CMVf
  12. H3160-2 - DD123
  13. H3180-1 - CMVf
  14. H3180-1 - DD123
  15. H3180-2 - CMVf
  16. H3180-2 - DD123
  17. H31116-2 - CMVf
  18. H31116-2 - DD123
  19. H31136-1 - CMVf
  20. H31136-1 - DD123





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