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Team:Paris/Addressing testing
From 2009.igem.org
iGEM > Paris > WetLab > Addressing
WetLab - Addressing the message
Global constructions :
Experiments ran :
| Column 1 | Column 2 | Column 3 | Column 4 |
| PCR :
pBAD: plate 2008
Oligo :O57 and O58 TM :
Oligo :O59 and O60 TM :
|
- |
- |
- |
| Verification on gel :
ok |
- |
- |
-
|
| Purification on gel :
ok |
-
|
- |
-
|
| Digestion:
ClyA Cterm S/P
| Digestion:
ClyA Cter-RFP Nter vector PSB1A3 E/X |
- |
- |
| Verification digestion:
ok | Verification digestion:
ok | - |
- |
| Ligation:
ClyA Cter-RFP Nter vector PSB1A3 | Ligation:
PBAD E/S ClyA Cter-RFP Nter PSB1A3 E/X (x2)
| Ligation:
- |
|
| Colony PCR :
ok | Colony PCR :
ok | Colony PCR :
ok |
|
| Miniprep:
clone : | Miniprep:
clone : | Miniprep:
clone 3 clone 5 |
|
| Sequencing :
ok | Sequencing :
ok | Sequencing :
ok |
-
|
| Stock glycerol:
ClyA CTer: S47(clone 3) and S48(clone 7) ClyA Nter: S72 RFP Cter: S55 (Clone 3) RFP Nter: S56 (Clone 3)
| Stock glycerol
Cly A(Cter)-(Nter)RFP: S72(clone 8) | Stock glycerol
pBAD ClyA RFP : S89 (clone 3) pBAD ClyA RFP: S90 (clone 5) |
|
Functional Testing:
PBAD ClyA RFP was transformed into Top10 bacteria in order to localize the fluorescence, we are supposed to have a superior fluorescence in the membrane.
PBAD ClyA RFP on PSB3T5 was transformed into Delta Tol bacteria , in this case we are supposed to see fluorescent vesicles went the medium contains 1 % arabinose , and to have no fluorescent on 1% glucose.
Export system
We finally thought that it won't be neccesary to overexpress the Tat system, nevertheless we have run a few experiments before starting to focus on others parts of the project.
Experiments ran :
| Column 1 | Column 2 | Column 3 | Column 4 |
| PCR :
TatABCE matrix : | - | - | -
|
| Verification on gel :
ok | - |
- |
- |
| Purification on gel :
ok | Purification on gel :
- |
- |
-
|
| Digestion:
none | Digestion:
none | Digestion:
none | Digestion:
none |
| Verification digestion:
none | Verification digestion:
none
| Verification digestion:
none | Verification digestion:
none
|
| Ligation:
none | Ligation:
none | Ligation:
none | Ligation:
none |
| Colony PCR :
none | Colony PCR :
none | Colony PCR :
none | Colony PCR :
none |
| Miniprep:
none | Miniprep:
none | Miniprep:
none | Miniprep:
none |
| Sequencing :
none | Sequencing :
none | Sequencing :
none | Sequencing :
none |
| Stock glycerol:
none | Stock glycerol
none | Stock glycerol
none | Stock glycerol
none |
