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Team:Paris Saclay/Notebook/August/20
From 2014.igem.org
(→Bacterial culture) |
(→CAD PCR) |
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Very small quantity of DNA | Very small quantity of DNA | ||
| - | 1.5 µl --> | + | 1.5 µl --> DMSO |
| - | 0.25 | + | 0.25 µl --> Gotaq |
| + | |||
| + | PCR cycle | ||
| + | |||
| + | 95° 2 min | ||
| + | 95° 30 sec | ||
| + | 58° 30 sec | ||
| + | 72° 1 min 45 | ||
| + | 72° 5 min | ||
| + | |||
| + | |||
| + | |||
| + | ====B - Construction of the fusion protein (color)===== | ||
| + | PCR with Gotaq | ||
| + | |||
| + | 22.75 µl --> H2O | ||
| + | |||
| + | 10 µl --> 5xbuffer | ||
| + | |||
| + | 1 µl --> dNTP | ||
| + | |||
| + | 4 µl --> MgCl2 | ||
| + | |||
| + | 5 µl iPS 83 (Primer) | ||
| + | |||
| + | 5 µl iPS 84 | ||
| + | |||
| + | 0.5 µl --> DNA | ||
| + | |||
| + | 1.5 µl --> DMSO | ||
| + | |||
| + | 0.25 µl --> Gotaq | ||
| + | |||
| + | |||
| + | PCR cycle | ||
| + | |||
| + | 95° 2 min | ||
| + | 95° 30 sec | ||
| + | 58° 30 sec | ||
| + | 72° 1 min 45 | ||
| + | 72° 5 min | ||
Revision as of 14:38, 20 August 2014
Contents |
Wednesday 20th August
D- Lemon scent
Bacterial culture
by Laetitia
Here are the results of the bacterial culture of yesterday
Bacteria with the pGEMTeasy without the DNA insert are blue and the bacteria with the pGEMTeasy and the DNA insert (PS, GS or LS gene) are white
Then, we have chosen 3 white colonies for each DNA insert and we have launched 3 corresponding liquid cultures :
- 5mL LB + ampiciline (1/1000e)
- 1 white colony
The 9 cultures are incubated at 37°C
by Melanie
Preculture 20ml of DH5a pPSI (apra 1/2000)
CAD PCR
Today we have receive our synthetic gene CAD (cinnamyl alcohol deshydrogenase) :)
so we do a PCR (with go taq):
23.25 µl --> H2O
10 µl --> 5xbuffer
1 µl --> dNTP
4 µl --> MgCl2
5 µl iPS 79 (Primer)
5 µl iPS 80
Very small quantity of DNA
1.5 µl --> DMSO
0.25 µl --> Gotaq
PCR cycle
95° 2 min 95° 30 sec 58° 30 sec 72° 1 min 45 72° 5 min
B - Construction of the fusion protein (color)=
PCR with Gotaq
22.75 µl --> H2O
10 µl --> 5xbuffer
1 µl --> dNTP
4 µl --> MgCl2
5 µl iPS 83 (Primer)
5 µl iPS 84
0.5 µl --> DNA
1.5 µl --> DMSO
0.25 µl --> Gotaq
PCR cycle
95° 2 min 95° 30 sec 58° 30 sec 72° 1 min 45 72° 5 min
