Team:Paris/14 August 2009
From 2009.igem.org
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==NoteBook== | ==NoteBook== | ||
- | + | {{Paris2009_Calendar}} | |
- | { | + | {{Paris2009_Calendar_Link|13_August_2009|15_August_2009}} |
- | + | <center> '''August 14th''' </center> | |
- | + | ||
- | + | ||
- | + | ||
- | + | ||
- | + | ||
<html> | <html> | ||
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</html> | </html> | ||
+ | ==Lab work== | ||
+ | <div class="charlotte"> | ||
+ | Digestion/Gel purification | ||
+ | </div> | ||
+ | <div class="experience"> | ||
- | + | Digestion of pSB1A3 w/: | |
+ | - EcoRI/PstI | ||
+ | - EcoRI/SpeI | ||
+ | - XbaI/PstI | ||
+ | ''Mix'' | ||
- | + | DNA : 10 uL | |
+ | |||
+ | 10X Buffer : 3 uL | ||
+ | |||
+ | Enz1 : 1 uL | ||
+ | |||
+ | Enz2 : 1uL | ||
+ | |||
+ | BSA : 0,5 uL | ||
+ | |||
+ | H2O : 14,5 uL | ||
- | |||
</div> | </div> | ||
- | <div | + | <div class="vicard"> |
+ | Digestion/Purification | ||
</div> | </div> | ||
- | <div | + | <div class="experience"> |
- | </ | + | <strong>Digestion of:</strong> |
+ | |||
+ | ClyA(A10) in X/P---->D30 | ||
+ | |||
+ | PSB2K3(P27) in X/P-->D31 | ||
+ | |||
+ | using: | ||
+ | |||
+ | H2O : 20,5µL | ||
+ | |||
+ | DNA : 20µL | ||
+ | |||
+ | BSA : 0,5µL | ||
+ | |||
+ | Buffer 2(x10) : 5µL | ||
+ | |||
+ | Xba : 2µL | ||
+ | |||
+ | Pst : 2µL | ||
+ | </div> | ||
+ | |||
+ | <div class="vicard"> | ||
+ | Ligation/Transformation | ||
+ | </div> | ||
+ | <div class="experience"> | ||
+ | Ligation of ClyA C-ter fusion cut in X/P (D31) in PSB1A3 cut in X/P (P25) | ||
+ | |||
+ | DO insert D31(ClyA C-ter fusion):2µg/mL (1/100) | ||
+ | DO vector P25(PSB1A3):0,60µg/mL (1/100) | ||
+ | |||
+ | Insert is two times smaller than the vector so we have to put 2times more insert. | ||
+ | |||
+ | <strong>x10 mix:</strong> | ||
+ | 1µL vector | ||
+ | |||
+ | (2*10)20µL insert | ||
+ | |||
+ | 2,5µL buffer 10x | ||
+ | |||
+ | 1µL ligase | ||
+ | |||
+ | 2µL H2O | ||
+ | |||
+ | <strong>x3 mix:</strong> | ||
+ | 1µL vector | ||
+ | |||
+ | (2x3)6µL insert | ||
+ | |||
+ | 1µL buffer 10X | ||
+ | |||
+ | 1µL ligase | ||
+ | |||
+ | 1µL H2O | ||
+ | |||
+ | <strong>negative control:</strong> | ||
+ | 1µL vector | ||
+ | |||
+ | 1µL buffer 10x | ||
+ | |||
+ | 1µL ligase | ||
+ | |||
+ | 7µL H2O | ||
+ | |||
+ | Then tranformation by Guillaume | ||
+ | </div> | ||
+ | |||
+ | |||
- | |||
- | |||
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</html> | </html> | ||
- | + | ==To do list== | |
- | + | ||
{| | {| | ||
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|- style="background: #bababa;text-align: center;" | |- style="background: #bababa;text-align: center;" | ||
|Sylvain | |Sylvain | ||
- | |New Glycerol stocks, OmpA-Linker : new PCR | + | |New Glycerol stocks, OmpA-Linker : new PCR, test the new clone miniprep (Charlotte) with purif |
|- style="background: #009900;text-align: center;" | |- style="background: #009900;text-align: center;" | ||
|Guillaume | |Guillaume | ||
| | | | ||
|} | |} | ||
+ | |||
+ | {{Paris2009_Calendar_Link|13_August_2009|15_August_2009}} |
Latest revision as of 13:21, 31 August 2009
NoteBook
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Lab work
Digestion/Gel purification
Digestion of pSB1A3 w/: - EcoRI/PstI - EcoRI/SpeI - XbaI/PstI
Mix
DNA : 10 uL
10X Buffer : 3 uL
Enz1 : 1 uL
Enz2 : 1uL
BSA : 0,5 uL
H2O : 14,5 uL
Digestion/Purification
Digestion of:
ClyA(A10) in X/P---->D30
PSB2K3(P27) in X/P-->D31
using:
H2O : 20,5µL
DNA : 20µL
BSA : 0,5µL
Buffer 2(x10) : 5µL
Xba : 2µL
Pst : 2µL
Ligation/Transformation
Ligation of ClyA C-ter fusion cut in X/P (D31) in PSB1A3 cut in X/P (P25)
DO insert D31(ClyA C-ter fusion):2µg/mL (1/100) DO vector P25(PSB1A3):0,60µg/mL (1/100)
Insert is two times smaller than the vector so we have to put 2times more insert.
x10 mix: 1µL vector
(2*10)20µL insert
2,5µL buffer 10x
1µL ligase
2µL H2O
x3 mix: 1µL vector
(2x3)6µL insert
1µL buffer 10X
1µL ligase
1µL H2O
negative control: 1µL vector
1µL buffer 10x
1µL ligase
7µL H2O
Then tranformation by Guillaume
To do list
Matricule | TODO |
Luc | |
Romain | |
Charlotte | |
Stoff | |
Chris | |
Lisa | |
Caroline | |
Souf | |
Vicard | |
Pierre | |
Sylvain | New Glycerol stocks, OmpA-Linker : new PCR, test the new clone miniprep (Charlotte) with purif |
Guillaume |