"
Team:Paris Saclay/Notebook/August/8
From 2013.igem.org
Notebook : August 8
Lab work
A - Aerobic/Anaerobic regulation system
((((((====Obtaining the PSB3K3 backbone plasmid====
1 - Electrophoresis of BBa_J004450 digested by EcoRI/Pst1 to check if the digestion was right
Damir, Nadia
| IMAGE |
|
Expected sizes :
- PSB3K3 : 2750kb
- GFP : 1069kb
We obtained fragments of the right size. Now we can purify the PSB3K3 plasmid.
2 - Gel purification of the PSB3K3 plasmid from BBa_J004450 digested by EcoRI/Pst1
2.1 - Migration of the remaining 45µL of BBa_J004450 digested by EcoRI/Pst1
Anaïs
|
|
Now we can do a gel purification of the highest band which contains the PSB3K3 plasmid.
2.2 - Electroelution of the highest band to extract the PSB3K3 plasmid from the gel
Nadia
Protocol : Electroelution
We let the plasmid precipitate during the night.))))))))))))))))))))))
Objective : obtaining Bba_K1155007
1 - Colony PCR of Bba_K115007 in DH5α
Anaïs
|
Tranformation of 08/07/13 works. we will make a PCR Colony. |
Colonie repiquée dans 10µL d'eau pour chaque tube ???????
Used quantities :
- DNA : 2µL
- Mix : (it was divided in 25 tubes for each promotor with 23µL of mix in each on)
- Oligo ... : 3.5µL
- Oligo ... : 3.5µL
- Buffer Dream Taq : 70µL
- dNTP : 28µL
- Dream Taq : 5µL
- H2O : 590µL
PCR Program :
2 - Electrophoresis to check the colony PCR products : Bba_K1155007
Anaïs, Damir
| 350px |
|
Expected size :
- Bba_K1155007 : 3583 bp
|
We obtain fragment at the right size for colonies 10, 14 and 15. We will extract BBa_K1155007. |
A - Aerobic/Anaerobic regulation system / B - PCB sensor system
Objective : obtaining FNR and BphR2 proteins
1 - Extraction of plasmid of BphR2, FNR, RBS-FNR
Damir
|
Transformation of 08/02/13 works. We will extract plasmids. |
Protocol : [http://www.mn-net.com/tabid/1452/default.aspx Gel purification ]
|
We lost our plasmids. We will do the Gibson assembly again. |
2 - Gel purification of RBS-BphR2 Part I
Nadia, XiaoJing
Protocol : [http://www.mn-net.com/tabid/1452/default.aspx Gel purification ]
|
We lost fragment. We will do the PCR again. |
