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Team:Lyon-INSA-ENS/Realisation/Week11
From 2011.igem.org
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<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| + | Nanodrop quantification of several midipreps :<br/> | ||
| + | p116 : 726.3 ng/µL<br/> | ||
| + | piG30 : 251.6 ng/µL<br/> | ||
| + | p115: 488.4 ng/µL<br/> | ||
| + | p10 : 276.2 ng/µL<br/> | ||
| + | p127 : 7673.5 ng/µL<br/><br/> | ||
| + | Start of solid cultures of S18 and S21 from the collection<br/><br/> | ||
| + | |||
| + | Digestion and electrophoresis of p115 (from S19 and NM522), p116(from S19 and NM522), p127(from S19 and NM522), p10(from NM522), p157 (from S19 and NM522), 2 clones each.<br/> <br/> | ||
| + | |||
| + | Plating of PHL1414 from the collection <br/> <br/> | ||
</p> | </p> | ||
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<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| + | Start of 5mL liquid cultures for transformations : PHL1414, MC4100, MC4100+pIG2<br/> | ||
| + | TSS Transformation and plating of PHL1414 with pIG2, MC4100 with R1 and R1+p150, MC4100+pIG2 with p150.<br/><br/> | ||
</p> | </p> | ||
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<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| + | Obtention of a few clones on all previous transformations except on MC4100+R1 ( too many clones, suspicious )<br/><br/> | ||
| + | |||
| + | TSS Transformation and plating of PHL1414 with pIG6 (Amp), MC4100 with pIG6+p150(Amp+Kan),R1(Cm), pIG26(Cm) and pIG26+p150(Cm+Kan). Negative controls : PHL1414 and MC4100 on every type of antibiotic previously used. <br/><br/> | ||
</p> | </p> | ||
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<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| + | The negative control for the transformations of MC4100 on Cm grew : start of several tests ( plating on solid medium with Cm, liquid cultures in LB+Cm) to identify the origin of the failure of the negative control.<br/> | ||
| + | Isolation of 4 individual clones on other antibiotic selctions.<br/><br/> | ||
| + | |||
| + | Plating of PHL1414+pIG6 and PHL1414+pIG2 on LB+Amp for TUdeflt collaboration <br/><br/> | ||
| + | |||
| + | Start of 2.5mL cultures for extraction and checking of the individual clones <br/><br/> | ||
| + | |||
| + | The tests for the failure of the negative transformation control showed that our starting MC4100 strain was resistant to Cm : all transformations using this strain are eliminated. <br/><br/> | ||
| + | |||
| + | Miniprep of PHL1414 + pIG2 and PHL1414+pIG6 ( 4 clones each ) | ||
</p> | </p> | ||
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<h6 style="text-align :left"> Friday </h6> | <h6 style="text-align :left"> Friday </h6> | ||
<p style=" line-height : 1.5em"> | <p style=" line-height : 1.5em"> | ||
| + | Digestion of the previous minipreps by E and electrophoresis : all clones are correct <br/> | ||
| + | Storage of clone 3 from PHL1414+pIG2 (S23) and clone 2 from PHL1414+pIG6(S24) <br/> | ||
| + | Plating of those same clones on LB+Amp medium.<br/><br/> | ||
| + | Start of flocculation tests :<br/> | ||
| + | -MC4100+pIG6 ( with and without cobalt ), MC4100+pÎG2 ( with and without cobalt ),in M63G+Amp at 30°C, 75rpm.<br/> | ||
</p> | </p> | ||
Revision as of 13:58, 2 September 2011
Week 11
From Monday the 29th of August to Friday the 2nd of September 2011
Monday
Nanodrop quantification of several midipreps :
p116 : 726.3 ng/µL
piG30 : 251.6 ng/µL
p115: 488.4 ng/µL
p10 : 276.2 ng/µL
p127 : 7673.5 ng/µL
Start of solid cultures of S18 and S21 from the collection
Digestion and electrophoresis of p115 (from S19 and NM522), p116(from S19 and NM522), p127(from S19 and NM522), p10(from NM522), p157 (from S19 and NM522), 2 clones each.
Plating of PHL1414 from the collection
Tuesday
Start of 5mL liquid cultures for transformations : PHL1414, MC4100, MC4100+pIG2
TSS Transformation and plating of PHL1414 with pIG2, MC4100 with R1 and R1+p150, MC4100+pIG2 with p150.
Wednesday
Obtention of a few clones on all previous transformations except on MC4100+R1 ( too many clones, suspicious )
TSS Transformation and plating of PHL1414 with pIG6 (Amp), MC4100 with pIG6+p150(Amp+Kan),R1(Cm), pIG26(Cm) and pIG26+p150(Cm+Kan). Negative controls : PHL1414 and MC4100 on every type of antibiotic previously used.
Thursday
The negative control for the transformations of MC4100 on Cm grew : start of several tests ( plating on solid medium with Cm, liquid cultures in LB+Cm) to identify the origin of the failure of the negative control.
Isolation of 4 individual clones on other antibiotic selctions.
Plating of PHL1414+pIG6 and PHL1414+pIG2 on LB+Amp for TUdeflt collaboration
Start of 2.5mL cultures for extraction and checking of the individual clones
The tests for the failure of the negative transformation control showed that our starting MC4100 strain was resistant to Cm : all transformations using this strain are eliminated.
Miniprep of PHL1414 + pIG2 and PHL1414+pIG6 ( 4 clones each )
Friday
Digestion of the previous minipreps by E and electrophoresis : all clones are correct
Storage of clone 3 from PHL1414+pIG2 (S23) and clone 2 from PHL1414+pIG6(S24)
Plating of those same clones on LB+Amp medium.
Start of flocculation tests :
-MC4100+pIG6 ( with and without cobalt ), MC4100+pÎG2 ( with and without cobalt ),in M63G+Amp at 30°C, 75rpm.
