IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/07/31

From OpenWetWare

Jump to: navigation, search
iGEM Project name 1 Main project page
Previous entry      Next entry

Biofilm Track

Picked two colonies (1 from each TSA plates preped by Jason and Melody) and placed in tube with 5ml TSB. for ON culture growth. (static condition, 37C, 20hours)

DspB Track

Marianne, Vicki

PCR gDNA (HK1561)

  • Resuspend primers in water to a concentration of 100uM
  • Primers:
  1. Upstream dspB His: 0.285mL water + 28.5nmol primer
  2. Upstream dspB: 0.312mL water + 31.2nmol primer
  3. Downstream dspB: 0.411mL water + 41.1nmol primer
  4. DspB fw rxn1: 0.290mL water + 29.0nmol primer
  5. DspB rev rxn1: 0.387mL water + 38.7nmol primer
  6. DspB fw rxn2: 0.342mL water + 34.2nmol primer
  7. DspB rev rxn2: 0.431mL water + 43.1nmol primer
  • Rehydrate Aggregatibacter actinomycetemcomitans: 250uL sdH2O into vial with 5ug of dried DNA. Vial in 37C for 1 hour.

Protocol: Common protocols for PCR

PCR Master mix
Reagent1x rxn volume (uL)Master Mix
5x rxn buffer5x1575
10mM dNTP2x1530
sdH2O8.8x15132
Phusion polymerase0.2x153
DNA5x1575
Total21315
  • Add 2uL of fw and rev primers to each PCR tube.
  • Elongation time: 40s
PCR Tubes
LabelContent
AdspB His 1
BdspB His 2
CdspB His 3
DdspB no His 1
EdspB no His 2
FdspB no His 3
GdspB rxn1 1
HdspB rxn1 2
IdspB rxn1 3
JdspB rxn2 1
KdspB rxn2 2
LdspB rxn2 3
MH2O control


PCR Cycles:

  • 98C @ 30s
  • Cycle 27x:
    • 98C @ 10 sec
    • 72C @ 30 sec
    • 72C @ 40 sec
    • 72C @ 10 min
  • 10C @ hold

Start: 1422
End: 1522

  • In 4C @ 1530

Vicki Ma 03:00, 3 August 2010 (EDT)


Personal tools