Team:Paris/18 August 2009
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==Lab work== | ==Lab work== | ||
+ | <div class="charlotte"> | ||
+ | PCR on colony w/ Soufifi | ||
+ | </div> | ||
+ | <div class="experience"> | ||
- | |||
+ | The ligation pLAC/RBS-TetR worked --> no clone on the negatif control and some clone on the "good" plates. | ||
+ | PCR on colony is required to check the validity of our clones. | ||
+ | |||
+ | |||
+ | ''Mix : total volume = 25uL'' | ||
+ | |||
+ | Super Mix 2X: 12,5 uL | ||
+ | VF2 (10uM): 0,5 uL | ||
+ | VR (10 uM): 0,5 uL | ||
+ | H20 : 6,5 uL | ||
+ | diluted bacteria : 5 uL | ||
+ | |||
+ | Tm = 55°C | ||
+ | Elongation time = 1 min | ||
- | |||
</div> | </div> | ||
- | <div | + | |
+ | |||
+ | |||
+ | <div class="charlotte"> | ||
+ | Digestion | ||
</div> | </div> | ||
- | <div | + | <div class="experience"> |
- | + | ||
+ | The ligation that I have set up yesterday (pSB1A3 [https://2009.igem.org/Team:Paris/Freezer_Digestion_Products#P16 pSB1A3] w/ D13 [https://2009.igem.org/Team:Paris/Freezer_Digestion_Products#D13 D13] or D14 [https://2009.igem.org/Team:Paris/Freezer_Digestion_Products#D14 D14] ) didn't work so I decided to redigest PCR products (A11 (RBS-ompA signal) and A13 (TolRII). | ||
+ | ''Mix : total volume = 30 uL'' | ||
+ | |||
+ | '''For A11:''' | ||
+ | |||
+ | DNA = 15 uL | ||
+ | |||
+ | 10X buffer: 3 uL | ||
+ | |||
+ | Enz1: 1 uL | ||
+ | |||
+ | Enz2: 1 uL | ||
+ | |||
+ | BSA: 0,5 uL | ||
+ | |||
+ | H2O: 9,5 uL | ||
+ | |||
+ | |||
+ | |||
+ | '''For A11:''' | ||
+ | |||
+ | DNA = 10 uL | ||
+ | 10X buffer: 3 uL | ||
+ | |||
+ | Enz1: 1 uL | ||
+ | |||
+ | Enz2: 1 uL | ||
+ | |||
+ | BSA: 0,5 uL | ||
+ | |||
+ | H2O: 14,5 uL | ||
+ | |||
+ | </div> | ||
==To do list== | ==To do list== | ||
edit please ^^ | edit please ^^ |
Revision as of 09:40, 18 August 2009
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NoteBook
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Brain work
edit please ^^
Lab work
PCR on colony w/ Soufifi
The ligation pLAC/RBS-TetR worked --> no clone on the negatif control and some clone on the "good" plates.
PCR on colony is required to check the validity of our clones.
Mix : total volume = 25uL
Super Mix 2X: 12,5 uL VF2 (10uM): 0,5 uL VR (10 uM): 0,5 uL H20 : 6,5 uL diluted bacteria : 5 uL
Tm = 55°C Elongation time = 1 min
Digestion
The ligation that I have set up yesterday (pSB1A3 pSB1A3 w/ D13 D13 or D14 D14 ) didn't work so I decided to redigest PCR products (A11 (RBS-ompA signal) and A13 (TolRII).
Mix : total volume = 30 uL
For A11:
DNA = 15 uL
10X buffer: 3 uL
Enz1: 1 uL
Enz2: 1 uL
BSA: 0,5 uL
H2O: 9,5 uL
For A11:
DNA = 10 uL 10X buffer: 3 uL
Enz1: 1 uL
Enz2: 1 uL
BSA: 0,5 uL
H2O: 14,5 uL
To do list
edit please ^^