IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/09/27

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Biofilm Track

Melody and Eric F.

Biofilm Protocol Day 4

  • 15 minute Crystal Violet Stain on plates EF09251020-1 to EF09251030-3

DspB Track

  • O/N culture for sonication taken out at 12pm; placed in 4C
  • Results: all tubes cloudy; J23100 is slightly red, rest is yellow colour
  • Made 1/100 dilutions for pre-dspB & 14 (Pcon + RBS) with C1 antibiotic
    • Put in 37C incubator at 12:15pm
    • Results: Lid closed too tight --> dspB = 0.081; 14 = 0.477
    • Final OD: pre-dspB = 1.15; 14 = 0.91
    • Centrifuge down into pellet
      • 1 mL into each microcentrifuge each time at 6000rcf for 5min each time (all 5mL)

Rafael's Method=

  • Mix rxn 1 with rxn 2 in rxn1 tube
  • Run in PCR machine:
    • 3 cycles:
      • 65C @ 10 min
      • 37C @ 20 min
  • Start: 8pm
  • End: 9:33pm

Ligation- Rafael's Method

Ligation
REAGENTS1 RXN VOLUME (uL)
Ligase1
T4 ligase buffer2
Insert3
Vector1
sdH2O8
Total15
  • Tubes: Raf
  • Content: backbone XP + insert (1) DspB
  • Incubate at room temperature for 20min
  • Store in -20C


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