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Team:Paris Saclay/Notebook/August/2
From 2013.igem.org
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==='''A - Aerobic/Anaerobic regulation system'''=== | ==='''A - Aerobic/Anaerobic regulation system'''=== | ||
| - | ===='''Objective : obtaining | + | ===='''Objective : obtaining BBa_K1155003'''==== |
| - | ===='''1 - Extraction of | + | ===='''1 - Extraction of BBa_K1155003 from DH5α'''==== |
Damir, Nadia | Damir, Nadia | ||
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{| | {| | ||
| style="border:1px solid black;padding:5px;background-color:#DE;" | | | style="border:1px solid black;padding:5px;background-color:#DE;" | | ||
| - | Tranformation from 07/30/13 works. We will extract | + | Tranformation from 07/30/13 works. We will extract BBa_K1155003 |
|} | |} | ||
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We mix our DNA in 50µL of H2O. | We mix our DNA in 50µL of H2O. | ||
| - | ===='''2 - Electrophoresis to check the extraction of | + | ===='''2 - Electrophoresis to check the extraction of BBa_K1155003'''==== |
Damir, Nadia | Damir, Nadia | ||
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| style="width:350px;border:1px solid black;vertical-align:top;" | | | style="width:350px;border:1px solid black;vertical-align:top;" | | ||
* Well 1 : 6µL of DNA Ladder | * Well 1 : 6µL of DNA Ladder | ||
| - | * Well 2 : 5µL of | + | * Well 2 : 5µL of BBa_K1155003 from clone 9+1µl of 6X loading dye |
| - | * Well 3 : 5µL of | + | * Well 3 : 5µL of BBa_K1155003 from clone 11+1µl of 6X loading dye |
| - | * Well 5 : 5µL of | + | * Well 5 : 5µL of BBa_K1155003 from clone 12+1µl of 6X loading dye |
* Gel : 0.8% | * Gel : 0.8% | ||
|} | |} | ||
Expected sizes : | Expected sizes : | ||
| - | * | + | * BBa_K1155003 : 2734 pb |
Estimated concentrations : | Estimated concentrations : | ||
* Clone 9 : 18ng/µL | * Clone 9 : 18ng/µL | ||
* Clone 11 :18ng/µL | * Clone 11 :18ng/µL | ||
| - | *Clone 12 : 18ng/µL | + | * Clone 12 : 18ng/µL |
{| | {| | ||
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|} | |} | ||
| - | ===='''Objective : obtaining | + | ===='''Objective : obtaining BBa_K1155007'''==== |
| - | ===='''1 - Digestion of | + | ===='''1 - Digestion of BBa_I732017 by EcoRI/SpeI to check sizes of fragments'''==== |
Damir, Nadia | Damir, Nadia | ||
Used quantities : | Used quantities : | ||
| - | * | + | * BBa_I732017 : 41µL |
* Buffer FD : 5µL | * Buffer FD : 5µL | ||
* EcoRI : 2µL | * EcoRI : 2µL | ||
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We let the digestion at 1h30 at 37°C. | We let the digestion at 1h30 at 37°C. | ||
| - | ===='''2 -Electrophoresis to check the digestion of | + | ===='''2 -Electrophoresis to check the digestion of BBa_I732017 by EcoRI/SpeI'''==== |
Damir, Nadia | Damir, Nadia | ||
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| style="width:350px;border:1px solid black;vertical-align:top;" | | | style="width:350px;border:1px solid black;vertical-align:top;" | | ||
* Well 1 : 6µL of DNA Ladder | * Well 1 : 6µL of DNA Ladder | ||
| - | * Well 2 et 3 : 5µL of | + | * Well 2 et 3 : 5µL of BBa_I732017 digested by EcoRI/SpeI+1µL 6X loading dye |
* Gel : 0.8% | * Gel : 0.8% | ||
|} | |} | ||
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{| | {| | ||
| style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | | style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | ||
| - | We don't obtain fragments at the right size. We will digest | + | We don't obtain fragments at the right size. We will digest BBa_KI732017 again. |
|} | |} | ||
Revision as of 09:30, 30 September 2013
Notebook : August 2
Lab work
A - Aerobic/Anaerobic regulation system
Objective : obtaining BBa_K1155003
1 - Extraction of BBa_K1155003 from DH5α
Damir, Nadia
|
Tranformation from 07/30/13 works. We will extract BBa_K1155003 |
Protocol : Hight copy plamid extraction
We used colony 9, 11 and 12. We mix our DNA in 50µL of H2O.
2 - Electrophoresis to check the extraction of BBa_K1155003
Damir, Nadia
| File:Ps.jpg|350px]] |
|
Expected sizes :
- BBa_K1155003 : 2734 pb
Estimated concentrations :
- Clone 9 : 18ng/µL
- Clone 11 :18ng/µL
- Clone 12 : 18ng/µL
|
We obtain fragment at the right size (Clone 12 starts 5 min after the others). The extraction was good. We will digest it. |
Objective : obtaining BBa_K1155007
1 - Digestion of BBa_I732017 by EcoRI/SpeI to check sizes of fragments
Damir, Nadia
Used quantities :
- BBa_I732017 : 41µL
- Buffer FD : 5µL
- EcoRI : 2µL
- SpeI : 2µL
We let the digestion at 1h30 at 37°C.
2 -Electrophoresis to check the digestion of BBa_I732017 by EcoRI/SpeI
Damir, Nadia
| [[]] |
|
Expected sizes :
- RBS-LacZ : 3093 bp
- PSB1A2 : 2079 bp
|
We don't obtain fragments at the right size. We will digest BBa_KI732017 again. |
A - Aerobic/Anaerobic regulation system / B - PCB sensor system
Objective : obtaining FNR and BphR2 proteins
1 - Transformation of BphR2, FNR and RBS-FNR in DH5α
XiaoJing
Protocol : Bacterial transformation
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