Team:Paris/August 4

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(New page: {{Paris/Calendar_Links|August 3|August 5}} == '''PCR Screening of the Ligation Transformants'''== Use of 8 clones of Ligation transformants n°L100 for screening PCR ===Protocol of s...)
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== '''PCR Screening of the Ligation Transformants'''==
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== '''PCR Screening of Ligation Transformants'''==
Use of 8 clones of Ligation transformants n°L100 for screening PCR  
Use of 8 clones of Ligation transformants n°L100 for screening PCR  
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===Results===
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===Results for L100===
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L100= D110 + D130 = '''RBS-tetR-ECFP-Ter''' [[Image:Part_icon_rbs.png]][[Image:Icon_coding.png]][[Image:Part_icon_rbs.png]][[Image:Part_icon_reporter.png]][[Image:Part_icon_terminator.png]]
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{| border="1"  
{| border="1"  
|align="center"|'''Band'''
|align="center"|'''Band'''
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|align="center"|n°1
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|align="center"| 1-->8 
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|align="center"|n°2
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|align="center"|n°3
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|align="center"|n°4
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|align="center"|n°5
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|align="center"|n°6
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|align="center"|n°7
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|align="center"|n°8
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|align="center"|'''Expected size'''
|align="center"|'''Expected size'''
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|colspan="8"| 1042 pb
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|align="center"|1042 pb
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|align="center"|'''Measured size'''
|align="center"|'''Measured size'''
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|align="center"|
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|align="center"|1100 pb
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|align="center"|
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|colspan="8"|[[Image: .jpg|thumb|'''Gel : L100(1-8)''']]
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|colspan="8"|[[Image:KR000101_1.jpg|thumb|'''Gel : L100(1-8)''']]
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|}
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==> '''Conclusion :''' we obtain a low quantity of PCR products, but at the good size.
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<br>So now, we know that we can amplify the clones that we want for L100 transformants bacteria.

Latest revision as of 15:22, 6 August 2008

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Contents

PCR Screening of Ligation Transformants

Use of 8 clones of Ligation transformants n°L100 for screening PCR


Protocol of screening PCR

  • Mix
Name Vol (µl) Concentration
Quick Load 25µl 2X
OligoF_VF2 (O18) 1µl 10µM
OligoR_VR (O19) 1µl 10µM
water 23µl


  • 50µl of Mix PCR by tube/clone
  • one toothpick of each clone's colony by tube
  • Program : Annealing 55°C - Time élongation 1'30" - Number cycle : 29


Conditions of electrophoresis

  • 10µl of ladder 1 kb
  • 20µl of screening PCR
  • migration ~30min at 100W on 0,8% gel


Results for L100

L100= D110 + D130 = RBS-tetR-ECFP-Ter Part icon rbs.pngIcon coding.pngPart icon rbs.pngPart icon reporter.pngPart icon terminator.png

Band 1-->8
Expected size 1042 pb
Measured size 1100 pb
Gel : L100(1-8)


==> Conclusion : we obtain a low quantity of PCR products, but at the good size.
So now, we know that we can amplify the clones that we want for L100 transformants bacteria.

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