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Team:Paris Saclay/Notebook/July/1
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(→1 - Digestion of PSB1C3 plasmid and PCR products : Pndh* by EcoRI/ PstI) |
(→2 - Ligation of PSB1C3 and Pfnr) |
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We let the digestion at 37°C during 3 hours. | We let the digestion at 37°C during 3 hours. | ||
| - | ===='''2 - Ligation of PSB1C3 and | + | ===='''2 - Ligation of PSB1C3 and Pndh*'''==== |
Sheng, Zhou | Sheng, Zhou | ||
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* Mix A : we mix our digestion mixes : | * Mix A : we mix our digestion mixes : | ||
** Digestion mix of PSB1C3 : 4µL | ** Digestion mix of PSB1C3 : 4µL | ||
| - | ** Digestion mix of | + | ** Digestion mix of Pndh* : 30µL |
** Buffer ligation : 2µL | ** Buffer ligation : 2µL | ||
** H2O : 14µL | ** H2O : 14µL | ||
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We let the ligation 1h30. | We let the ligation 1h30. | ||
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==='''B - PCB sensor system'''=== | ==='''B - PCB sensor system'''=== | ||
Revision as of 13:47, 29 September 2013
Contents |
Notebook : July 1
Lab work
A - Aerobic/Anaerobic regulation system
Objective : obtaining Bba_K1155000
1 - Digestion of PSB1C3 plasmid and PCR products : Pndh* by EcoRI/ PstI
Zhou
Used quantities :
- PSB1C3 :
- Plasmid : 4µL
- EcoRI FD : 0.5µL
- PstI FD : 0.5µL
- Buffer FD : 0.8µL
- H2O : 2.2µL
- Pndh* :
- Pndh* : 20µL
- EcoRI FD : 0.75µL
- PstI FD : 0.75µL
- Buffer FD : 3µL
- H2O : 5.5µL
We let the digestion at 37°C during 3 hours.
2 - Ligation of PSB1C3 and Pndh*
Sheng, Zhou
Used quantities :
- Mix A : we mix our digestion mixes :
- Digestion mix of PSB1C3 : 4µL
- Digestion mix of Pndh* : 30µL
- Buffer ligation : 2µL
- H2O : 14µL
- Then, we denature EcoRI/SpeI used for the digestion by ethanol precipitation.
Protocol : Ethanol precipitation
We used 50µL of DNA. At the end, we dilute our DNA in 20µL of H2O.
- Finally we did the ligation mix :
- Buffer ligation : 2µL
- Ligase : 1µL
- DNA : 2µL
- H2O : 15µL
We let the ligation 1h30.
B - PCB sensor system
Objective : obtaining BphR2 protein
1 - Design of oligos for amplification of BphR2 gene of Pseudomonas pseudoalcaligenes, Pseudomonas oleovorans
Abdou, Sheng, Zhou
We used software gene manager to find the correct oligopeptides for amplification of BphR2 genes in Pseudomonas pseudoalcaligenes, Pseudomonas oleovorans.
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