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Team:Paris Saclay/Notebook/August/28
From 2014.igem.org
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(→Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position) |
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| + | 1-5: pPS5 (pJBEI+CAD) digested by HindIII | ||
| + | 6: pPS1 (pJBEI+Apra) | ||
| + | 7-8: ladder | ||
====Electrophoresis of the same 5 PCR pooled==== | ====Electrophoresis of the same 5 PCR pooled==== | ||
Revision as of 10:32, 29 August 2014
Contents |
Thursday 28th August
Lab Work
B - Construction of the fusion protein (color)
Transformation of DH5a by PSB1C3+chromoprotein
by Mélanie
--> a compléter
Addition of adenines at the ends of PCR fragment of chromoprotein
by Laetitia
| components | volumes |
|---|---|
| H2O | 1 μl |
| Gotaq buffer 5X | 1µl |
| dATP 1mM | 2 µl |
| Chromoprotein gene fragment (30ng/µL) | 5µL |
| Gotaq polymerase | 1µl |
1h- 70°C
Ligation of the PCR fragment of chromoprotein + AAA with pGEMTeasy
by Laetitia
| components | volumes |
|---|---|
| 2X ligation buffer T4 DNA ligase | 10 μl |
| pGEMTeasy | 1µl |
| Ligation mix | 7 µl |
| Ligase | 2µL |
4h - 16°C
D - lemon scent
Electrophoresis of 5 digestions of pPS5 by HindIII to check if the insert of CAD is in the right position
by Hoang Vu
1-5: pPS5 (pJBEI+CAD) digested by HindIII
6: pPS1 (pJBEI+Apra)
7-8: ladder
Electrophoresis of the same 5 PCR pooled
by Laetitia
Gel agarose 0,8% - 100V
The totality of the 5 PCR have been used. Then, we cut the lane corresponding of the Limonene synthase gene on a UV table.
Purification of the PCR product of Limonene synthase (BBa762100)
by Laetitia We used the kit PCR-Clean-UP to extract the DNA from the agarose gel. Elution volume : 15µl
