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Team:Paris Bettencourt/Notebook/Trojan Horse/Tuesday 10th September.html
From 2013.igem.org
Trojan Horse
ASDF10th September
Place your twit here
Experiment Silencing Cm (try to reduce the noise, aka the variability)
- Launch O/N culture of producing phages cells ; centrifuge ; take supernatant and filter it.
-diluting 100x O/N of MGZ1, F+ -pACYc in LB-Cm (25 ug/ul)
- At OD 0,7 :. Put 900 uL of cells in each tube. Add in tubes respectively : nothing, 100uL phages of litmusGFP-LacZ1, litmusGFP-Cm1 & litmusGFP-Cm2
-3 replicates for each condition. Plate each Dilution 2 times to limit the variability
Non infected cells (1,2,3)
Cells infected with anti lacZ (1,2,3)
Cells infected with anti CmA (1,2,3)
Cells infected with antiCmB (1,2,3
- Incubate 45 minutes at 37°C
- Serial Dilute every tube until 10-5. Plate 100ul for each tube according to this dilution table
|
LB |
Cm 25 ug/ul |
Cm 250ug/ul |
Cm 500 ug/ul |
|
|
non infected 1 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
|
non infected 2 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
|
non infected 3 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
10^-5/4 |
|
sRNA lacZ 1 |
10^-4 |
10^-4 |
10^-4/2 |
10^-2/1 |
|
sRNA lacZ 2 |
10^-4 |
10^-4 |
10^-4/2 |
10^-2/1 |
|
sRNA lacZ 3 |
10^-4 |
10^-4 |
10^-4/2 |
10^-2/1 |
|
sRNA Cm A 1 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
|
sRNA Cm A 2 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
|
sRNA Cm A 3 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
|
sRNA Cm B 1 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
|
sRNA Cm B 2 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
|
sRNA Cm B 3 |
10^-4/3 |
10^-4/3 |
10^-3 |
10^-2/1 |
