IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/09/14

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dspB Track

Miniprep

  • Resuspend with 100uL of H2O
  • Nanodrop:
    • 450 HRA: 594.45ng/uL
    • 450NCD: 1290.25ng/uL
    • 450 HRC: 682.85ng/uL
    • 450 HRB: 835.7ng/uL
    • 450 NCC: 1288.3ng/uL

Restriction Digest

Restriction Digest Supermix
REAGENTS1 RXN VOLUME (uL)
Buffer 22
BSA0.2
DNA10
ddH205.8
Total20

Enzymes: Use EcoRI and SpeI (1uL each)
DNA: 1uL of RFP chloraphenicol backbone, and 5uL miniprepped DNA
Digest for 2 hours

Gel verification on RD

  • Protocol: biobrick "digest" protocol

Gel orientation:

Gel orientation
NCCHRCNCD100bp ladderHRAHRBRFP1kb ladder

Machine conditions: 0.5x TBE buffer, 100V, 45min Results:
[[Image:|200px]]

Transformations (HC)

  • Protocol: SOP; used competent cell #1
    • Add 5uL of ligation mixes
    • Plates put in 37C incubator at 11:46pm