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Team:Paris Saclay/Notebook/August/1
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CarolineMir (Talk | contribs) (→1 - Gel purification of PCR products : BphR2 Part I, BphR2, Part II, FNR Part I, FNR Part II, RBS-FNR Part I, PSB1C3) |
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===='''Objective : obtaining FNR and BphR2 proteins'''==== | ===='''Objective : obtaining FNR and BphR2 proteins'''==== | ||
| - | ===='''1 - Gel purification of PCR products : BphR2 Part I, BphR2 | + | ===='''1 - Gel purification of PCR products : BphR2 Part I, BphR2 Part II, FNR Part I, FNR Part II, RBS-FNR Part I PSB1C3 '''==== |
Xavier | Xavier | ||
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** BphR2 Part I : 1µL | ** BphR2 Part I : 1µL | ||
** BphR2 Part II : 1µL | ** BphR2 Part II : 1µL | ||
| - | ** Gibson mix : 15µL | + | ** Gibson mix : 15µL |
** H2O : 1µL | ** H2O : 1µL | ||
| Line 45: | Line 45: | ||
** FNR Part I : 1µL | ** FNR Part I : 1µL | ||
** FNR Part II : 1µL | ** FNR Part II : 1µL | ||
| - | ** Gisbon mix : 15µL | + | ** Gisbon mix : 15µL |
** H2O : 1µL | ** H2O : 1µL | ||
| Line 52: | Line 52: | ||
** RBS-FNR Part I : 1µL | ** RBS-FNR Part I : 1µL | ||
** FNR Part II : 1µL | ** FNR Part II : 1µL | ||
| - | ** Gibson mix : 15µL | + | ** Gibson mix : 15µL |
** H2O : 1µL | ** H2O : 1µL | ||
| Line 68: | Line 68: | ||
** DNA : 0.25µL | ** DNA : 0.25µL | ||
** dNTP : 1µL | ** dNTP : 1µL | ||
| - | ** Phusion : | + | ** Phusion : 5µL |
** H2O : 36.5µL | ** H2O : 36.5µL | ||
Revision as of 17:10, 27 September 2013
Notebook : August 1
Lab work
A - Aerobic/Anaerobic regulation system / B - PCB sensing system
Objective : obtaining FNR and BphR2 proteins
1 - Gel purification of PCR products : BphR2 Part I, BphR2 Part II, FNR Part I, FNR Part II, RBS-FNR Part I PSB1C3
Xavier
Protocol : [http://www.mn-net.com/tabid/1452/default.aspx Gel purification ]
Nanodrop :
- BphR2 Part I : 44 ng/µL
- BphR2 Part II : 64 ng/µL
- FNR Part I : 147 ng/µL
- FNR Part II : 140 ng/µL
- RBS-FNR Part I : 167 ng/µL
- PSB1C3 : 159 ng/µL
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The PCR was good. Now we will do the Gibson assembly. |
2 - Gibson assembly.
Abdou, Xiaojing
Used quantities :
- BphR2 :
- PSB1C3 : 2µL
- BphR2 Part I : 1µL
- BphR2 Part II : 1µL
- Gibson mix : 15µL
- H2O : 1µL
- FNR :
- PSB1C3 : 2µL
- FNR Part I : 1µL
- FNR Part II : 1µL
- Gisbon mix : 15µL
- H2O : 1µL
- RBS-FNR :
- PSB1C3 : 2µL
- RBS-FNR Part I : 1µL
- FNR Part II : 1µL
- Gibson mix : 15µL
- H2O : 1µL
We let these mix at 50°C during 1h.
3 - PCR of : RBS-BphR2 Part I
Abdou
Used quantities :
- Oligo 54F : 1µL
- Oligo 55R : 1µL
- Buffer phusion : 5µL
- DNA : 0.25µL
- dNTP : 1µL
- Phusion : 5µL
- H2O : 36.5µL
PCR Program :
3 - Electrophoresis of the PCR products : RBS-BphR2 Part I
Xavier, XiaoJing
| File:Ps.jpg|350px]] |
|
Expected sizes :
- RBS-BphR2 Part I : ....
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We obtain fragments at the right size. We will purify it. |
