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Team:Lyon-INSA-ENS/Realisation/Week12
From 2011.igem.org
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<FONT COLOR="red"> <b>Transformations and controls for future tests</b> </FONT> <br/><br/> | <FONT COLOR="red"> <b>Transformations and controls for future tests</b> </FONT> <br/><br/> | ||
| + | <b>Miniprep</b> of three individual clones of MC4100/piG6 and MC4100/piG2 isolated on 09/02 in order to chek the presence of the right plasmid.<br/><br/><br/> | ||
| + | </p> | ||
| + | <p style=" line-height : 1.5em"> | ||
| + | <FONT COLOR="purple"> <b>Results of Flocculation Test and New Test</b> </FONT> <br/><br/> | ||
| + | Crystal Violet coloration for the series of 09/02.<br/><br/> | ||
| + | |||
| + | Start of a fourth test in M63G medium with :<br/> | ||
| + | -MC4100/piG6 (AmpR) without Co and with Co 10µM (negative controls)<br/> | ||
| + | -MC4100/piG2 (AmpR) without Co and with Co 10µM, 25µM, 50µM, 100µM.<br/> | ||
| + | (three repetitions)<br/><br/> | ||
| + | |||
| + | These liquid cultures are let for two days at 30°C and low stirring (75).<br/><br/><br/> | ||
| + | </p> | ||
<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| - | + | <FONT COLOR="purple"> <b>Adherence Test – pRcn-csgBAEFG Characterization</b> </FONT> <br/><br/> | |
| + | Start of three replica of 24 well plates in M63G medium + Amp with MC4100/piG6 without Co and with Co 10µM<br/> (negative control) and MC4100/piG2 without Co and with Co 10µM, 25µM, 50µM, 100µM.<br/> | ||
| + | Incubation at 30°C for 48h.<br/><br/> | ||
| + | |||
| - | + | Revealing of the well plates started on 09/02.<br/> | |
| - | + | Measuring OD600 and Crystal Violet coloration.<br/> | |
| - | + | ||
| - | |||
<br/> <br/> | <br/> <br/> | ||
| - | |||
</p> | </p> | ||
Revision as of 11:58, 15 September 2011
Week 12
From Monday the 5th of September to Friday the 9th of September 2011
Monday
Transformations and controls for future tests
Miniprep of three individual clones of MC4100/piG6 and MC4100/piG2 isolated on 09/02 in order to chek the presence of the right plasmid.
Results of Flocculation Test and New Test
Crystal Violet coloration for the series of 09/02.
Start of a fourth test in M63G medium with :
-MC4100/piG6 (AmpR) without Co and with Co 10µM (negative controls)
-MC4100/piG2 (AmpR) without Co and with Co 10µM, 25µM, 50µM, 100µM.
(three repetitions)
These liquid cultures are let for two days at 30°C and low stirring (75).
Adherence Test – pRcn-csgBAEFG Characterization
Start of three replica of 24 well plates in M63G medium + Amp with MC4100/piG6 without Co and with Co 10µM
(negative control) and MC4100/piG2 without Co and with Co 10µM, 25µM, 50µM, 100µM.
Incubation at 30°C for 48h.
Revealing of the well plates started on 09/02.
Measuring OD600 and Crystal Violet coloration.
Tuesday
TSS transformation of MC4100 with pIG27 (Cm) or pIG16 (Amp).
CFA tests
Digestion and electrophoresis of the previously extracted DNA of MC4100+pIG6 clones and MC4100+pIG2 clones. Comparison made with the parts put in the collection.
Wednesday
Results of tuesday transformations : no clone on the negative controls, several on the tests. Isolation of 4 clones per transformation in solid and liquid culture.
Pouring of Amp plates.
Revelation of the 2 plates from september 5th.
Thursday
E+P digestion and electrophoresis of all the isolated clones for the following transformations : MC4100+pIG27, MC4100 + pIG16, MC4100 + rcn-ompR234.
All clones are correct for pIG27 and pIG16. No clone is correct for rcn-ompR234 -> detection of an error in the ligation.
Storage of clone 1 for pIG27 and pIG16. Plating of those same clones on LB+antibiotic (Cm and Amp respectively ).
Friday
Nanodrop quantification of the previous pIG27 miniprep : 48.0 ng/µL.
Preparation of tubes for sequencing (30µL at approximately 50-100 ng/µL). The pIG27 miniprep is used as such. pIG25 is diluted from 5µL of the collection (371.4 ng/µL) with 25µL water.
