Team:Paris/August 21
From 2008.igem.org
(Difference between revisions)
(→Screening of the cloning of E0240 and FlhDC+promotor) |
(→Electrophoresis) |
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|'''sample''' | |'''sample''' | ||
- | |1 kb DNA ladder | + | |1 kb<br> DNA ladder |
- | | | + | |control + |
- | | | + | |control - |
|colspan="13"|S159.1 | |colspan="13"|S159.1 | ||
- | |100 bp DNA ladder | + | |100 bp<br>DNA ladder |
|- | |- | ||
|'''colonie n°''' | |'''colonie n°''' | ||
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|- | |- | ||
|'''sample''' | |'''sample''' | ||
- | |1 kb DNA ladder | + | |1 kb<br>DNA ladder |
- | | | + | |control + |
- | | | + | |control - |
|colspan="13"|S161.1 | |colspan="13"|S161.1 | ||
- | |100 bp DNA ladder | + | |100 bp<br>DNA ladder |
|- | |- | ||
|'''colonie n°''' | |'''colonie n°''' | ||
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<br> | <br> | ||
- | '''Results''': | + | '''Results''': |
- | *The clone of E0240 (S159.1) always have several bands amplified by PCR. It might contain different plasmids. | + | *The clone of E0240 (S159.1) always have several bands amplified by PCR. It might contain different plasmids. |
- | *The clone of FlhDC+promotor (S161.1) don't have the correct size band. It also doesn't have the insert in the plasmid. | + | *The clone of FlhDC+promotor (S161.1) don't have the correct size band. It also doesn't have the insert in the plasmid. |
='''Construction for synchronization'''= | ='''Construction for synchronization'''= |
Revision as of 13:55, 6 September 2008
Cloning of FlhB promoterProtocol
Resuspension of 1 colony E.coli K12 strain MG 1655 in 100µl of water.
1µl of dNTP
Result
Construction for FIFOAim : Construction of pFlgA - YFP tripart (+/- LVA) "pFlgA-RBS-YFP-dbl ter" (pFlgA-E0430/E0432) DigestionDigestion
Gel Extraction
Measurement of the concentration of D166 & D167 purifiedProtocol (it's same that for Miniprep)
Ligation
Screening of the cloning of E0240 and FlhDC+promotorWe obtained colonies isolated with the dilution 1/100; 13 clones were analysed by PCR PCR screeningreaction mixture (25 µL)
PCR screening programm
Electrophoresis
Results: *The clone of E0240 (S159.1) always have several bands amplified by PCR. It might contain different plasmids. *The clone of FlhDC+promotor (S161.1) don't have the correct size band. It also doesn't have the insert in the plasmid. Construction for synchronizationLigations
Promoter characterization plasmidsLigations from digestions from 20thTop 10 cells were used
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