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Team:Bordeaux/NotebookAugust27
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- RLP: 429,9 ng/μl => 2,33 μl + 7,67μl H2O<br> | - RLP: 429,9 ng/μl => 2,33 μl + 7,67μl H2O<br> | ||
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Digestion of pSB1C3<br> | Digestion of pSB1C3<br> | ||
| - | Dilution DNA: pSB1C3: 224,2 ng/μl | + | Dilution DNA: pSB1C3: 224,2 ng/μl => 4,46μl + 5,54 H2O<br> |
Test with 1,5μl, 1μl, 0,5μl and 0μl of enzyme<br> | Test with 1,5μl, 1μl, 0,5μl and 0μl of enzyme<br> | ||
Gel | Gel | ||
Latest revision as of 02:35, 18 October 2014
27th August
Molecular biology
-
3 digestions (ELP, SLP and RLP)
We want remove the stop codon (with Nhe1, Spe1 and dephosphorylation) and recover our insert (ELp, SLP and RLP)
Dilution DNA :- ELP : 328,5 ng/μl => 3μl + 7μl H2O
- SLP : 266,4 ng/μl => 3,75μl + 6,25μl H2O
- RLP: 429,9 ng/μl => 2,33 μl + 7,67μl H2O
Buffer 4 10X 5μl Enzyme 1 1μl Enzyme 2 1μl DNA 10μl H2O qsp 50μl
=> Gel - Ligation (assembly)
-
Digestion of pSB1C3
Dilution DNA: pSB1C3: 224,2 ng/μl => 4,46μl + 5,54 H2O
Test with 1,5μl, 1μl, 0,5μl and 0μl of enzyme
Gel -
Ligation (pSB1C3)
=> 30min at 45°C and transformationPlasmid 4μl Insert 4μ Buffer 1μl Taq ligase 0,5μl H2O qsp 10μl
