Difference between revisions of "Team:Paris Saclay/Notebook/August/29"

(→‎PCR of the plasmid extraction from clonies 4,5,9,12 containing FRB sequence in pJET plasmid)
(→‎PCR of the plasmid extraction from clonies 4,5,9,12 containing FRB sequence in pJET plasmid)
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|Tm
 
|Tm
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|°C
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|52°C
 
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|t
 
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!Expected band size (bp)
 
!Expected band size (bp)
 
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|dCas9 NM - GFP 10 - pSB1C3
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|FRB in pJET
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|3688
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|474
 
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|}
  
 
[[File:T--Paris Saclay--160826 gel ligation fragm1 et 2.jpg.png|400px|thumb|center|Result of the migration]]
 
[[File:T--Paris Saclay--160826 gel ligation fragm1 et 2.jpg.png|400px|thumb|center|Result of the migration]]
  
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GEL GEL GEL
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====NanoDrop Measurements====
 +
''By Mahnaz''
 +
 
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{| class="wikitable"
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!Sample
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!Concentration (ng/µL)
 +
|-
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|PCR fragment GFP 11 clone 6<div id="PCR fragment GFP 11 - pSB1C3 clones 6"></div>
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|187.23
 +
|-
 +
|PCR fragment GFP 11 clone 8<div id="PCR fragment GFP 11 - pSB1C3 clones 8"></div>
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|156.85
 +
|-
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|PCR fragment FRB clone 4<div id="PCR fragment FRB clone 4"></div>
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|75.67
 +
|-
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|PCR fragment FRB clone 9<div id="PCR fragment FRB clone 9"></div>
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|246.41
 +
|-
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|PCR fragment GFP 1.9<div id="PCR fragment GFP 1.9"></div>
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|22.06
 +
|-
 +
|}

Revision as of 08:17, 16 September 2016

PCR sur colonie

PCR of the plasmid extraction from clonies 4,5,9,12 containing FRB sequence in pJET plasmid

By Mahnaz

  • 2.5 µL DreamTaq Buffer
  • 0.5 µL of dNTPs (10mM)
  • 1 µL of each primer (10µM) primer pJET R and F
  • 0.13 μl of DreamTaq Pol
  • 20 µl H2O

PCR was performed as follow:

Step Temperature Time
Initial denaturation 95°C 3 min
30 cycles 95°C 30 sec
52 30 sec
72°C 1 min
Final Extension 72°C 7min
Hold 4°C \infinity\ 
Primers used were:
Matrix Clones containing dCas9 NM - GFP 10 in pSB1C3
Primers pJET R and F
Tm 52°C
t 1 min 30

After amplification, 3 µL of each PCR products and 5 µL of DNA ladder were placed in wells and migrated at 100V during 30 min.

PCR products expected were :

PCR products Expected band size (bp)
FRB in pJET 474
Result of the migration

NanoDrop Measurements

By Mahnaz

Sample Concentration (ng/µL)
PCR fragment GFP 11 clone 6
 187.23
PCR fragment GFP 11 clone 8
 156.85
PCR fragment FRB clone 4
 75.67
PCR fragment FRB clone 9
 246.41
PCR fragment GFP 1.9
 22.06