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Team:Paris Liliane Bettencourt/Notebook/2011/09/05/
From 2011.igem.org
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Dilution of overnight cultures : YC164 and YC227 (SinOp Design) . <br> | Dilution of overnight cultures : YC164 and YC227 (SinOp Design) . <br> | ||
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| + | YC227 has a hyperspank promoter (inducible by IPTG) which regulates a GFP gene. It also produces phosphorilated KinA.<br> | ||
| + | YC164 Contains SpoA which can be indirectly phosphorilated by KinA. Phosphorilated SpoA indirectly activates a gene which contains GPF and SinI. SinI is a signal involved in biofilm shaped growth.<br> | ||
Two well slides : 1-control (YC164) 2-Mix (both) | Two well slides : 1-control (YC164) 2-Mix (both) | ||
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=== Observation === | === Observation === | ||
Revision as of 12:57, 7 September 2011
Contents |
Cyrille
Miniprep
Miniprep of 2 clones of TetO array in pSB1C3, 2 clones of TetO array in pHM3 Miniprep of Mathias: S21 S82 1 2 3 4; S75 S67 and S75
With good yeilds (>500ng)
Gel Extraction
Gel extraction of pHM3 Digested in EP and pVeg YFP TetR D EP and DXP.
Ligation
Ligation of pHM3 Digested EP with pVeg-YFP-TetR D. EP with ratio 1:1 1:10 1:100 Ligation pHM3 TetO D. SP with pVeg-YFP-TetR D. SP
Transformation
Transformation of the ligation product + ComS ligated in pSB1C3 + YFP TetR BB ligated in pSB1C3
Contol of the comptent cells
Two tests of transformation was done with the B7 plasmid in TurboCell competent cells.
The rest of the tube is placed in 500µL of LB, incubated for an hour and then plated.
Hovannes-Baptiste
Preparation of slides
Dilution of overnight cultures : YC164 and YC227 (SinOp Design) .
YC227 has a hyperspank promoter (inducible by IPTG) which regulates a GFP gene. It also produces phosphorilated KinA.
YC164 Contains SpoA which can be indirectly phosphorilated by KinA. Phosphorilated SpoA indirectly activates a gene which contains GPF and SinI. SinI is a signal involved in biofilm shaped growth.
Two well slides : 1-control (YC164) 2-Mix (both)
Observation
-37°C Microscopy-
YC164 grows fast
YC227 does not grow at all
YC164 does not change its state
We can see some GFP after an overnight
No significant results
