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Wisconsin-Madison/29 June 2009
From 2009.igem.org
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June 29, 2009
Ex 16:
MiniPrep: of P2 colonies A-C, ProU colonies A-3, NudF colonies A-3, GFP+ProU
Inoculate Cultures: YhfR, BB (P1)
U U U F F F B* B* B*UG L U F B* UG
Left of ladder-cut, right of ladder-uncut
Expected sizes: F:2700bp, U:2350bp, B*:2150bp, UG:2650bp
Ex 17: GFP Regulation by ProU Promoter
Experimental Design:
0.0-1.0M in increments of 0.2 concentrations of (4 wells of each concentration)
1. Wild Type E.Coli
2. GFP only
3. ProU + GFP
(Plan to include the SAM synthase (METK) to boost gsSDMT function, and eventually getting rid of the GFP and evaluating solely on growth)
4. ProU + GFP + gsSDMT
5. ProU + GFP + gsSDMT + MetK
6. Salt Free LB – start (2 wells of each concentration)
7. Water (2 wells of each concentration)
1. Grow four different cultures till OD600=0.4 to 0.6
2. Induce cultures with various concentrations of salt in the plate reader
3. Let the plate reader shake and incubate overnight
4. Take OD and excitation/emission value at 501/511nm during the entire time course
Test 4:
Conc of salt (M): 0.0 / 0.2 / 0.4 / 0.6 / 0.8 / 1.0
DH10B
GFP only
ProU+GFP
Salt Free LB
Water
0D 600
.401 GFP+ProU
.425 GFP
.415 DH10 B
- Incubator overnight (37 C)
Ex 10: Cyanobacteria
Cyano antibiotic test
Made 10:1 dilutions of 6803 to achieve 110 mL cultures, doubled the volume of transformed cells, made freezer stocks of extremely dark green 7942, tested antibiotics and other things with 7942 - NP
