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Team:Paris Saclay/Notebook/August/6
From 2014.igem.org
Contents |
Wednesday 6th August
Lab work
The chassis coli Odor free
Final stock of MG1655
by Romain
2 final stocks of the MG1655 made with 1ml of strain and 250µl glycerol 87%, in -80°C.
Salicylate Inducible Suppressing System
Plasmid DNA Purification
by Eugene and Fabio
- BBa_K1372000 Cl.1
- BBa_K1372000 Cl.2
From Liquid Culture made the 5th August
Digestion
by Eugene and Fabio
In order to place the BBa_B0015 just after the BBa_K1372000, we will start the [http://parts.igem.org/Help:Assembly/3A_Assembly Assembly procedure] again.
- BioBrick BBa_K1372000 (Salicylate promoter + NahR + RNA suppressor) as Part A
- BioBrick BBa_B0015 (Double terminator) as Part B
The enzymes used were:
- EcoRI and SpeI for BBa_K1372000
- EcoRI and XbaI for BBa_B0015
TODO: Illustration of the process
BioBrick Assembly - Digest Reaction Protocol
Segregate Process by Electrophoresis
by Eugene and Fabio
- BBa_B0015 digested by EcoRI and XbaI
- BBa_K1372000 digested by EcoRI and SpeI
BioBrick Assembly - Segregate Process Protocol
Lemon scent
PCR Clean-up
by Pierre & Sean
BioBricks used: BBa_K762100 and BBa_K517003 (PCR perfomed on the 4th August)
PCR digestion
by Pierre & Sean
Protocol
For each BioBrick (Bba_K517003 and BBa_K762100)mix the following into a microcentrifuge tube.
| component | volume |
|---|---|
| H2O | 5.5μl |
| PacI | .5μl |
| Fast Digest buffer | 4μl |
| Cleaned-up PCR content | 30μl |
Then bain-marie at 37°C for one hour. Finally conduct PCR cleanup for both BioBricks, with 15µl of NE buffer.
We acidentally used PvuI instead of PacI. - SC 6/8/14
PCR pCola plasmide (Geranyl Synthase)
by melanie
amplification of geranyl Synthase
PCR protocol: 98° --> 2min
5 PCR cycle:
| time | 10sec | 20sec | 45sec |
|---|---|---|---|
| temperature | 98° | (depending on the tube - see the gradient) | 72° |
25 PCR cycle
| time | 10sec | 20sec | 45sec |
|---|---|---|---|
| temperature | 98° | 72° | 72° |
and last step: 72° during 10min
but we don't have any results
Gel electrophoresis
by Sean,Pierre & Melanie
Samples used:
- BBa_K762100 (2µl)
- BBa_K517003 (2µl)
- p cola(12µl)
Bibliography
By Hoang Vu
Still working on my mission: find more informations about limonene, pinene and citral A and B.I got now a couple of very interessant reviews about limonene and pinene.
Photo of the Day
Members present:
- Instructors and advisors: Alice.
- Students: Eugene, Fabio, Hoang Vu, Juliette, Melanie, Pierre, Romain, Sean and Terry.
