Team:Northwestern/08 26

Notebook

Friday, August 26th

Tasks:

Michelle

  • Gel extracted gRNA and signal sequences
    • Used IBI protocol (kit from Leonard Lab)
    • Nanodrop results: all under 10 ng/uL, 260/280 above 1.9, 260/230 around 0.5 — discarded
  • Reran signal sequence PCR (to add homology)
    • 1 uL template
    • 1 uL fwd primer 10uM
    • 1 uL rev primer 10uM
    • 1 uL DMSO
    • 21 uL nfH2O
    • 25 uL OneTaq Master Mix
  • Ran 2 gels on the signal sequence PCR products

Tasfia

  • DpnI digested Cas9-Lrz-SS PCR product
  • Made more 10X TAE
  • Registered all the ChemEs and Jordan for iGEM Jamboree
  • DpnI digested Tet-Lrz-GFP for two hours and ran a gel screen on it (with 1 uL of our remaining SybrGreen)
    • Only primer dimers result from the Tet-Lrz-GFP PCR

Tyler

  • Gibson Assembly of gRNA-mRFP
    • gRNA-mRFP Assembly
      • 0.31 µL insert
      • 0.4 µL backbone
      • 4.29 µL water
      • 5 µL Gibson mix
    • Negative Control
      • 0.4 µL backbone
      • 4.6 µL water
      • 5 µL Gibson Mix
    • Quantities transformed (heat shocked 30s, 200µL SOC)
      • 3 µL gRNA-mRFP assembly product
      • 2 µL negative control
      • 2µL positive Gibson kit control
      • 1µL positive transformation control

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