Team:Paris/August 8
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{{Paris/Calendar_Links|August 7|August 9}} | {{Paris/Calendar_Links|August 7|August 9}} | ||
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==== '''PCR verification/Analyse''' ==== | ==== '''PCR verification/Analyse''' ==== | ||
- | [[Image:KR000129.jpg | + | [[Image:KR000129.jpg|thumb|Analysis of PCR product]] |
''After the PCR :'' | ''After the PCR :'' | ||
* 3µl have been analysed by electrophoresis | * 3µl have been analysed by electrophoresis | ||
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|style="background: #cbff7B"|<center> 1400 pb</center> | |style="background: #cbff7B"|<center> 1400 pb</center> | ||
|} | |} | ||
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+ | ==> '''Conclusion :''' we observed the size expected for the PCR products, but not for FlhDC gene.<br> We hypothesis that its value of amplification for this gene it's low as we can't visualize it. So we try to continue to experiments to know if there is something inside or not this tube. | ||
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|1.84 | |1.84 | ||
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=== Digestion of PCR products === | === Digestion of PCR products === | ||
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==== Purification/Analysis ==== | ==== Purification/Analysis ==== | ||
- | + | [[Image:KR000142.jpg|thumb|Analysis of PCR product]] | |
'''Electrophoresis''' | '''Electrophoresis''' | ||
- | * ladder : 10µl ladder 1 kb | + | * ladder : 10µl ladder 1 kb |
* samples : 3µl of insert + 2µl of Loading Dye | * samples : 3µl of insert + 2µl of Loading Dye | ||
- | * Conditions : migration 30min at 100W, on a '''1 | + | * Conditions : migration 30min at 100W, on a '''1''' agarose gel. |
* Results : | * Results : | ||
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|'''Name''' | |'''Name''' | ||
|'''Promotor''' | |'''Promotor''' | ||
- | |||
|align="center"|'''Band''' | |align="center"|'''Band''' | ||
|align="center"|'''Expected size''' | |align="center"|'''Expected size''' | ||
|align="center"|'''Measured size''' | |align="center"|'''Measured size''' | ||
|- style="text-align: center;" | |- style="text-align: center;" | ||
- | + | |D139 | |
- | + | |FlhDC gene | |
- | + | | | |
- | + | ||
|style="background: #cbff7B"|<center> 972 pb</center> | |style="background: #cbff7B"|<center> 972 pb</center> | ||
|align="center"| pb | |align="center"| pb | ||
|- style="text-align: center;" | |- style="text-align: center;" | ||
- | + | |D140 | |
- | + | |OmpR gene | |
- | + | | | |
- | + | ||
|style="background: #cbff7B"|<center> 762 pb</center> | |style="background: #cbff7B"|<center> 762 pb</center> | ||
- | + | | pb | |
|- style="text-align: center;" | |- style="text-align: center;" | ||
- | + | |D141 | |
- | + | |EnvZ gene | |
- | + | | | |
- | + | ||
|style="background: #cbff7B"|<center> 1421 pb</center> | |style="background: #cbff7B"|<center> 1421 pb</center> | ||
- | + | |pb | |
|- style="text-align: center;" | |- style="text-align: center;" | ||
- | + | |D142 | |
|align="center"| -vector- | |align="center"| -vector- | ||
|align="center"| | |align="center"| |
Revision as of 14:39, 11 August 2008
Minipreps : Plasmid extractionExtraction of pSB3K3 et E0240 in pSB1A2 plasmid from overnight bacteria culture using the QIAspin Miniprep Kit (QIAGEN) by QIACube.
Amplification of Genes of interest (OmpR, EnvZ, FlhDC)We performed PCR on to amplify the sequence in order to have enough amount of DNA to carry out the following of our experiments.
PCR amplificationProtocol
For each samples, 1 µl dNTP
PCR verification/AnalyseAfter the PCR :
==> Conclusion : we observed the size expected for the PCR products, but not for FlhDC gene.
Digestion of PCR productsProtocol :
Purification/AnalysisElectrophoresis
Transformation results
PCR Screening of Ligation TransformantsUse of 8 clones of Ligation transformants for screening PCR
Protocol of screening PCR
Conditions of electrophoresis
Results of electrophoresis
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