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Team:Northwestern/Notebook
From 2010.igem.org
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6/14-18/10 (Boot Camp)
DNA extracted from kit
Part: [1]
6/22/10
We poured our Kan, Amp, Tet, Kan+Amp, Kan+Tet, Amp+Tet antibiotic agar plates.
Antibiotic Concentrations
- Amp: 50mg/500ml
- Kan: 31.97mg/500ml
- Tet: 6mg/500ml
6/23/10
Resuspended 12O and 6G DNA from the iGEM Kit and transformed them into our competent cells.
6G = r0010(inducible promoter) 12O = e0840(rbs30-gfp-2xterm)
6/24/10
Ran a gel of our PCR product to make sure that our taq polymerase master mix was working.
Selected single 12O and 6G colonies and transferred them to LB and incubated for 18 hours.
Let 1 12O and 1 6G plate incubate at RT. Let 1 12O and 1 6G plate incubate at 37°C. Intended to observe lawn growth.
6/25/10
Low DNA yield; Plan to redo DNA extraction from Kit
Found pink colonies in 12O plates. Determined to be contaminations. Plates were thrown out.
6/28/10
Kit to Stock
1-12O (r0010: inducible promoter)
1-6G (e0840: rbs30-gfp-2xterm)
3-20M (K124017: Bacteriophage Lysis Cassette S105, R, and Rz)
Plates were incubated at 37°C overnight (starting at 2:30 pm).
6/29/10
Selected single 1-12O, 1-6G, 3-20M colonies and grew them in Amp-LB broth for 18 hours (starting at 4:15 pm).
6/30/10
Poured 2 Sleeves of Kan and Amp plates respectively
Met with Dr. Russin (BIF)
Sent emails regarding biofilm and biofilm imaging: Aaron Packman, Wei Zhang, Kimberly Grey
Most likely going to use water immersion Leica (Confocal Microscope) + low MP agar
Red colonies reappear on plates with 1-12O; Fluorescent Microscopy reveals negligible activity; still unsure what red colonies are.
Miniprepped 1-12O, 3-20M, 1-6G: nanodrop -> mostly below 20ng, 1 above 100ng; stored in -20°C
Re-miniprepped after growing in liquid LB for 3 hours; nanodrop -> around 50ng, 2 around 100ng; stored in -20°C
Transferred overnight cultures into new 5ml LB broths. Incubated overnight in an angled shaker (starting from 4:20pm)
Plated 25μl, 100μl, and 200μl (added LB up to 200μL) of the KR 1-12O overnight culture. Intend to measure the thickness of E.Coli lawn.
