IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 Arsenic Bioremediation/2010/08/19
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Made glycerol stock of LamBF2
.9 ML 50% glycerol, .9 mL LamBF2 overnight.
Incubate at 37 for 15°C min. Inactivate at 80°C for 20 min.
Incubate at room temperature for 10 min, then inactivate at 80°C for 20 min.
50 μL cells, 2 μL ligation product. 10 min on ice, 45 sec at 42°C, 2 min on ice. then add 1 mL SOC, recover for 1 hour at 37°C shaking.
It's hard to see, but there are very faint bands around 1.3 Kb which is the expected size for ArsB
Made 5 mL Cultures (2 colonies from each plate section)of the following samples:
ArsRSDM2-B0015-pSB1T3 plate had questionable looking colonies (hence re-doing yesterday's procedure today) but I picked what looked like colonies anyways.
No growth on pArsRF1R2 2 or pArsRF1R2 2' plates.
Plated 300 μL transformation product on Tet plate
ArsB50: 4.8 ng/μL ArsB53: 6.1 ng/μL ArsB56: 6.3 ng/μL
Francis Miniprepped ArsRSDM2 and LamBF2 overnights, both at 250 ng/μL.