Difference between revisions of "Team:Paris Saclay/Notebook/July/6"

(Lab work)
(Lab work)
 
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{{Team:Paris_Saclay/notebook_header}}
 
{{Team:Paris_Saclay/notebook_header}}
 
=Wednesday 6<sup>th</sup> July=
 
=Wednesday 6<sup>th</sup> July=
==Lab work==
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<!--Rédigé par Terrence, à relire -->
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====Preparation of LB solid and liquid stock====
 
====Preparation of LB solid and liquid stock====
 
''By Léa, Naiane, Laetitia''
 
''By Léa, Naiane, Laetitia''
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- 2L of LB liquid : 20g/L powder LB + 2L water μQ<br/>
 
- 2L of LB liquid : 20g/L powder LB + 2L water μQ<br/>
 
- 1L of LB solid : 1L of LB liquid +15g/L of Agar<br/>
 
- 1L of LB solid : 1L of LB liquid +15g/L of Agar<br/>
The all was put in the autoclave for sterilization with the help of Sylvain.
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The solutions were put in autoclave for sterilization with the help of Sylvain.
 
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===Bringing DNA closer===
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==== ====
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''By ''
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===Biobrick characterization===
 
===Biobrick characterization===
==== ====
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==== DNA extraction of K1372001 clone 1 and clone 2====
''By ''
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''By Mathilde''
  
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Plasmids DNA were extracted following the [[Team:Paris_Saclay/Experiments#PlasmidExtraction|usual]] protocol except that isopropanol was used instead of 100% ethanol and that pellets were dried on the lab bench top instead of with the speedvac. The extractions were kept at -20°C.
  
 
===Visualization===
 
===Visualization===
====Screen PCF for GBlock ====
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==== gBlocks PCR screening ====
 
''By Caroline''
 
''By Caroline''
  
<!--TODO :  A relire ABSOLUMENT problème de lecture et de traduction-->
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A PCR screening was carried out on colonies of bacteria transformed with the gBlocks (6 clones for each except pPS16_004 for which it was only 2 clones). We use the Taq [[Team:Paris_Saclay/Experiments#taqPCR|usual]] protocol. We used puc19 [[Team:Paris_Saclay/Experiments#primers|primers]] 1151_pheoR and 1152_pheoF.
"Megamix" were prepared for 34 tubes with 70μL of 90xTaq buffer + 70 μL of <!-- dumpamM --> + 42 μL MgCl<SUB>2</SUB> 25mM + 35 μL primer 1 + 35 μL primer 2 + 3.5 μL of Taq + 409.5 μL of water (till 700 μL completion, 35  μL corresponding to the 1 μL of liquid culture for each.)<br/>
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19 μL of mage was put into PCR's tubes then 1 μL of culture were added for each Gblock 1.1 1.2 2.1 2.2 3.1 and 3.2 and their 6 clones. (2 clones for 2.2).
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1 μL of each culture were added for each gblock pPS16_001, pPS16_002, pPS16_003, pPS16_004, pPS16_005, pPS16_006 and pPS16_007.
  
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==== pPS16_004, pPS16_007 and the pPS16_007 clone 6 (selected on 01/07/2016) streaks ====
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''By Léa, Caroline and Laetitia''
  
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The transformations carried out the 30/06/2016 were streaked again on LB Petri dishes containing 50µg/ml of amplicilin on which X-Gal/IPTG was spread on (white/blue screen). The Petri dishes were incubated ON at 37°C.
  
 
{{Team:Paris_Saclay/notebook_footer}}
 
{{Team:Paris_Saclay/notebook_footer}}

Latest revision as of 14:12, 9 October 2016

Wednesday 6th July

Preparation of LB solid and liquid stock

By Léa, Naiane, Laetitia

- 2L of LB liquid : 20g/L powder LB + 2L water μQ
- 1L of LB solid : 1L of LB liquid +15g/L of Agar
The solutions were put in autoclave for sterilization with the help of Sylvain.

Biobrick characterization

DNA extraction of K1372001 clone 1 and clone 2

By Mathilde

Plasmids DNA were extracted following the usual protocol except that isopropanol was used instead of 100% ethanol and that pellets were dried on the lab bench top instead of with the speedvac. The extractions were kept at -20°C.

Visualization

gBlocks PCR screening

By Caroline

A PCR screening was carried out on colonies of bacteria transformed with the gBlocks (6 clones for each except pPS16_004 for which it was only 2 clones). We use the Taq usual protocol. We used puc19 primers 1151_pheoR and 1152_pheoF.

1 μL of each culture were added for each gblock pPS16_001, pPS16_002, pPS16_003, pPS16_004, pPS16_005, pPS16_006 and pPS16_007.

pPS16_004, pPS16_007 and the pPS16_007 clone 6 (selected on 01/07/2016) streaks

By Léa, Caroline and Laetitia

The transformations carried out the 30/06/2016 were streaked again on LB Petri dishes containing 50µg/ml of amplicilin on which X-Gal/IPTG was spread on (white/blue screen). The Petri dishes were incubated ON at 37°C.