"
Team:Paris Saclay/Notebook/August/12
From 2013.igem.org
(Difference between revisions)
| Line 78: | Line 78: | ||
===='''Objective : obtaining FNR and BphR2 proteins (Gibson assembly)'''==== | ===='''Objective : obtaining FNR and BphR2 proteins (Gibson assembly)'''==== | ||
| - | ===='''1 - Electrophoresis of PCR products : RBS-BphR2 | + | ===='''1 - Electrophoresis of PCR products : RBS-BphR2 Part I, BphR2 Part I, BphR2 Part II, RBS-FNR Part I, FNR Part I and FNR Part II '''==== |
Damir | Damir | ||
| Line 86: | Line 86: | ||
| style="width:350px;border:1px solid black;vertical-align:top;" | | | style="width:350px;border:1px solid black;vertical-align:top;" | | ||
* Well 1 : 6µL DNA Ladder | * Well 1 : 6µL DNA Ladder | ||
| - | * Well 2 : 5µL RBS-BphR2 | + | * Well 2 : 5µL RBS-BphR2 Part I+1µl of 6X loading dye |
| - | * Well 3 : 5µL BphR2 | + | * Well 3 : 5µL BphR2 Part II+1µl of 6X loading dye |
| - | * Well 4 : 5µL FNR | + | * Well 4 : 5µL FNR Part I+1µl of 6X loading dye |
| - | * Well 5 : 5µL FNR | + | * Well 5 : 5µL FNR Part II+1µl of 6X loading dye |
| - | * Well 6 : 5µL RBS-FNR | + | * Well 6 : 5µL RBS-FNR Part I+1µl of 6X loading dye |
| - | * Well 4 : 5µL BphR2 | + | * Well 4 : 5µL BphR2 Part II+1µl of 6X loading dye |
* Gel : 0.8% | * Gel : 0.8% | ||
|} | |} | ||
Expected size | Expected size | ||
| - | * RBS-BphR2 | + | * RBS-BphR2 Part I : 197 kb |
| - | * BphR2 | + | * BphR2 Part II : 790 kb |
| - | * FNR | + | * FNR Part I : 597 kb |
| - | * FNR | + | * FNR Part II : 200 kb |
| - | * RBS-FNR | + | * RBS-FNR Part I : 615 kb |
| - | * BphR2 | + | * BphR2 Part I : 178 kb |
{| | {| | ||
| style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | | style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | ||
| - | We can't see FNR Part I, FNR | + | We can't see FNR Part I, FNR Part II and BphR2 Part i fragments at the good size. We will make the PCR again. We obtain RBS-BphR2 Part I, BphR2 Part II, RBS-FNR Part I frangments at the right size thanks to the PCR. We will purify it. |
|} | |} | ||
{{Team:Paris_Saclay/incl_fin}} | {{Team:Paris_Saclay/incl_fin}} | ||
Revision as of 21:08, 10 September 2013
Notebook : August 12
Lab work
A - Aerobic/Anaerobic regulation system
Objective : obtaining ...
1 - Digestion of Bba_K1155000 by SpeI/PstI, Bba_K1155007 and Bba_K1155003 by XBaI/PstI
Anaïs, Nadia, XiaoJing
Used quantities :
- Bba_K1155000 :
- Buffer FD : 5µL
- H2O : 38µL
- DNA : 5µL
- SpeI FD : 1µL
- PstI FD : 1µL
- Bba_K1155007 :
- Buffer FD : 5µL
- H2O : 23µL
- DNA : 20µL
- XBal FD : 1µL
- PstI FD : 1µL
- Bba_K1155003 :
- Buffer FD : 5µL
- H2O : 33µL
- DNA : 10µL
- XBal FD : 1µL
- PstI FD : 1µL
2 - Electrophoresis to check the digestion of Bba_K1155000 by SpeI/PstI, Bba_K1155007 and Bba_K1155003 by XBalI/PstI
Nadia
| File:Psdigestion12.jpg|350px]] |
|
Expected sizes :
- Pfnr : ...
- RBS_LacZ_Term : 3500 kb
- RBS_AmilCP_Term : ...
- PSB1C3 : ...
|
We can't see any band for Bba_K1155000 digestion. The digestion failed. We will do it again. We obtain RBS-LacZ-Term and RBS-AmilCP-Term fragments. The digestion was good. We will purify it. |
3 - Digestion of Bba_K1155000 by SpeI/PstI
Anaïs, Nadia
Used quantities :
- Buffer FD : 5µL
- H2O : 38µL
- DNA : 5µL
- SpeI FD : 1µL
- PstI FD : 1µL
A - Aerobic/Anaerobic regulation system / B - PCB sensor system
Objective : obtaining FNR and BphR2 proteins (Gibson assembly)
1 - Electrophoresis of PCR products : RBS-BphR2 Part I, BphR2 Part I, BphR2 Part II, RBS-FNR Part I, FNR Part I and FNR Part II
Damir
| [[]] |
|
Expected size
- RBS-BphR2 Part I : 197 kb
- BphR2 Part II : 790 kb
- FNR Part I : 597 kb
- FNR Part II : 200 kb
- RBS-FNR Part I : 615 kb
- BphR2 Part I : 178 kb
|
We can't see FNR Part I, FNR Part II and BphR2 Part i fragments at the good size. We will make the PCR again. We obtain RBS-BphR2 Part I, BphR2 Part II, RBS-FNR Part I frangments at the right size thanks to the PCR. We will purify it. |
