Team:Paris Saclay/Notebook/September/3
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We made a classic exttraction of plasmids from the liquid cultures. | We made a classic exttraction of plasmids from the liquid cultures. | ||
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''by Mélanie'' | ''by Mélanie'' | ||
Revision as of 09:13, 14 October 2014
Contents |
Wednesday 3rd September
LabWork
Construction of the fusion protein
We made a classic exttraction of plasmids from the liquid cultures.
Check by electrophoresis if it worked and send it for sequencing.
Lemon scent
by Mélanie
PCR LS
Electrophoresis of the PCR made Yesterday
well 2-3 = PCR with Dream taq or Vent taq
(well 4-5 = checking of the pps5 Extraction)
The PCR have success so I use the PCR clean up kit to purify it
Electrophoresis after purification
Digestion
In order to clone LS in pPSI, i Digest it by PacI
component | volume |
---|---|
H2O | 3μl |
buffer | 1μl |
PacI | 1μl |
PCR purify | 5μl |
Ligation
We already have some pPSI digested and dephosphorelated
So I do a ligation:
component | volume |
---|---|
H2O | 5μl |
buffer | 2μl |
ligase | 1μl |
pPSI | 2μl |
LS PCR | 10μl |
2 hours at room temperature and over night at 4°
pPS5
Plasmid exctraction using the kit (picture is here)
digestion by SalI
component | volume |
---|---|
H2O | 11μl |
buffer | 5μl |
SalI | 2μl |
pPS5 | 30μl |
Electrophoresis
We saw that we have something strang with our plasmid but we wil check it tomorrow with a good ladder (the ladder here was normally on the well 1)
pPS3 and pPS4
We digest the plasmid by HindIII to direct our insert
component | volume |
---|---|
H2O | 6μl |
buffer | 1μl |
HindIII | 1μl |
pPS3/4 | 2μl |
well 1 = ladder well 2-3-4 = pPS3 Well 5-6-7 = pPS4
results are very strange
Photo of the Day
[http://iramis.cea.fr/llb/Phocea/Pisp/visu.php?id=35&uid=%20alexei.grinbaum Alexei Grinbaum] , one of those few we have interviewed during this month of september.
To learn more about the ethic aspect of our project, just click here