"
Team:Yale/Notebook/Week4
From 2011.igem.org
Notebook: Week 4
Monday
- Made glycerol stock of Canada cells.
- Made plates 1mM IPTG + Amp
- Made 1 L LB, completed 1/40 dilution of ZeAFP (added Amp), shook for 3 hrs, checked OD until .78 OD reached
- Divided 1 L into 6 separate 100 mL solutions, half with IPTG, induced for 5 hours
- Centrifuged 4 100 mL solutions, saved supernatant/pellet for 2 (frozen at -20); for the other 2, we completed the following protocol:
1) Washed with 50 mL water
2) Resuspend in 100 mL ice-cold water
3) Divided 1ml aliquots into 8 eppendorf tubes (half induced half uninduced)
4) Added 100 and 10ul to IPTG and non IPTG plates (4 total) and grew at room temperature. Froze eppendorft tubes. Tomorrow we will unfreeze them for 6 hours on ice, then 1 hour at 4C, then replate to compare colony growth - essentially a survival assay after frozen : before frozen number of viable cells (this is obviously just a test assay)
- Plated 6 samples of the remaining 2 100 mL solutions, using 10 and 100 microliters on IPTG, LB, Amp plates and non-IPTG, LB, Amp plates and stored 1 of each concentration in 0 degrees, 4 degrees, and room temperature
Tuesday - Finished and presented project defense presentation to Sackler Institute and our advisers. - Made 3L worth of amp+iptg plates for large assay - Obtained TCA protein precipitation protocol. - Bacterial colonies are not growing yet - we will give more time b/c they were at room temperature previously.
Wednesday
Plan:
Accomplished:
Thursday
Plan:
Accomplished:
Friday
Plan:
Accomplished:
