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

From OpenWetWare

Jump to: navigation, search
Karmella's BioBrick Cloning Main project page
Previous entry      Next entry

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 = 2668, 252
Empty PLrigid = 2668, 95
4. DT004 = 2656, 432
Empty PLrigid4 = 2656, 275
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.
  2. DT002 - success.
  3. DT003 - success.
  4. DT004 - success.


  • Next steps:
    • Sequence verify all clones (extra precaution; linker part is small)



Sequencing order

  • 1-4 - Ben
  • 5-16 - Cas-tone
  1. DBN007-1 - VF2 - good
  2. DBN007-1 - VR - good, pSB1A3 terminator appears to be deleted
  3. DBN007-2 - VF2 - good
  4. DBN007-2 - VR - good, pSB1A3 terminator appears to be deleted
  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





Personal tools