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Team:Paris Saclay/Notebook/August/18
From 2014.igem.org
(→B - Construction of the fusion protein (color)) |
(→Ligation of PCR products inside pGEMTeasy) |
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|pGEMT easy(50ng) | |pGEMT easy(50ng) | ||
|1 μl | |1 μl | ||
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|DNA insert (GS or LS or PS PCR products) | |DNA insert (GS or LS or PS PCR products) | ||
Revision as of 16:31, 20 August 2014
Contents |
Monday 18th August
Lab work
B - Construction of the fusion protein (color)
PCR of the chromoprotein
27 µl H2O
10µl 5X phusion buffer
1µl dNTP 10mM
1 µl Primer (iPS83 and iPS84)
small amount of DNA
1,5µl DMSO
0,5µl Phusion
PCR cycle:
98° --> 30 sec
98° --> 10 sec
58° --> 30 sec
72° --> 45 sec
72°1 --> 10 min
D - Lemon Scent
PCR of BBa_K517003
By Hoang Vu and Eugène
We realized a PCR to amplify the β-pinene synthase gene.
| components | volumes |
|---|---|
| DNA of GS | 1 μl |
| green goTaq buffer 5X | 10 μl |
| GoTaq enzyme | 0.5 µl |
| H2O | 29.5 µl |
| MgCl2 | 4 µl |
| dNTP | 1µl |
| IPS68bis | 2 µl |
| IPS69bis | 2 µl |
PCR of BBa_K762100
by Sean
Five tubes were prepared. Protocol was modified to match the one used for BBa_K517003 and p cola (PCR conducted on 8th August).
Primers used: iPS66 and iPS67.
PCR of pCola
by Laetitia and Melanie
We realized a PCR to amplify the geraniol synthase gene.
Five tubes were prepared. Protocol was modified to match the one used for pCola (PCR conducted on 8th August).
Primers used : iPS91bis and iPS82
Electrophoresis of the 3 PCR
(image)
Ligation of PCR products inside pGEMTeasy
by Laetitia
We used the 3 previously PCR products to perform 3 reaction of ligation. The DNA insert used for each was : PS-6, LS-1 and GS-5
| components | volumes |
|---|---|
| (2X)T4 ligase buffer | 5μL |
| pGEMT easy(50ng) | 1 μl |
| DNA insert (GS or LS or PS PCR products) | 3μL |
| Ligase | 1 µl |
16°C until afternoon and 4°C during the night
