Team:Paris Saclay/Notebook/August/27
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====Ligation==== | ====Ligation==== | ||
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==== Transformation of odor free E. coli with plasmids coding Fluo Protein ==== | ==== Transformation of odor free E. coli with plasmids coding Fluo Protein ==== |
Revision as of 14:56, 28 August 2014
Contents |
Wednesday 27th August
Lab Work
B - Construction of the fusion protein (color)
PCR clean-up of colonies containing pGEMTeasy+chromoprotein
by Sean
Three colonies were selected from the gel prepared earlier by Laetitia, namely #3,#4, and #6. The three gel samples underwent PCR clean-up.
Gel electrophoresis of PCR clean-up results containing pGEMTeasy+chromoprotein
by Sean
PCR clean-up was performed ealier today. 1µl of each result was used for the electrophoresis. Nothing appeared under UV.
Ligation
by Sean
Transformation of odor free E. coli with plasmids coding Fluo Protein
by Terry
Our fusion protein was not expressed in pGEMTeasy (no color in the culture), so, if our system do not work at all, I'm preparing FP (Fluorescent Protein) for our lemon.
6 plamids coding FP have been transformed in Odor Free :
- pEYFP
- pGFP
- pESFP
- pRFP+
- pCFP+
- pYFP+
D - lemon scent
Plasmide extraction
By Mélanie
Extraction of pPS5 with the phenol protocol (as previously described)
PCR of Limonene synthase (BBa762100)
by Mélanie
5 tubes to do a gradient
components | volumes |
---|---|
H2O | 29.75μl |
Gotaq buffer | 10µl |
dNTP 10mM | 1µl |
iPS67 | 1µM |
iPS66 | 1µM |
DNA | 1µl |
MgCl2 | 4µl |
Gotaq | 1.25µl |
step | temperature (°C) | time |
---|---|---|
1 | 95 | 2min |
2 | 95 | 30 sec |
3 | 49-55 (gradient) | 30sec |
4 | 72 | 2min |
5 | 72 | 5min |
Electrophoresis of pPS3 and pPS4
by Terry
Result from yesterday 's extraction.
Alice, donne moi la photo.