!Clones containing corrected dCas9 ST - GFP 11 in pSB1C3 (pPS16_017)
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|Primers
|Primers
Line 71:
Line 71:
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GEL
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[[File:T--Paris Saclay--Gel111111.png|400px|thumb|center|Result of the migration]]
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====Colony PCR of 16 clones containing FRB - GFP 11 - GFP 1.9 in pSB1C3 (pPS16_022)====
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''By Maxence, Mahnaz & Coline''
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A colony PCR was done for 16 clones from the 20th September. For that purpose, 16 clones were screened and used for the usual [[Team:Paris_Saclay/Experiments##Polymerase_chain_reaction|protocol]] of Colony PCR. The same colonies were also plated on Petri dish containing solid LB + 30 µg/mL Cm and liquid cultures (LB + 30 µg/mL Cm) were made from these clones. Both petri dishes and liquid cultures were grown at 37°C.
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For each clones contained in 20 μl water, 5.13 μL of the following mix were added :
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* 2.5 µL DreamTaq Buffer
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* 0.5 µL of dNTPs (10mM)
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* 1 µL of each primer mix (10µM)
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* 0.13 μl of DreamTaq Pol
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PCR was performed as follow:
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{| class="wikitable"
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!Step
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!Temperature
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!Time
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|Initial denaturation
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|95°C
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|3 min
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|rowspan="3"|30 cycles
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|95°C
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|30 sec
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|48.4°C
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|30 sec
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|72°C
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|1min 30sec
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|Final Extension
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|72°C
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|7 min
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|Hold
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|4°C
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|$\infty$
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|}
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[[Team:Paris_Saclay/Experiments#primers|Primers]] used were:
