Team:Paris Saclay/Notebook/August/14
From 2013.igem.org
Line 76: | Line 76: | ||
Expected sizes : | Expected sizes : | ||
+ | *Pfnr in PSB1C3 : ... | ||
+ | *Nar K in PSB1C3, NarG in PSB1C3, NirB in PSB1C3 : ... | ||
*PSB1C3 : 2070kb | *PSB1C3 : 2070kb | ||
*NarK, NarG, NirB : 200kb | *NarK, NarG, NirB : 200kb | ||
- | *Pfnr : | + | *Pfnr : ... |
- | + | {| | |
- | + | | style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | |
+ | Well 1 : we have done one digestion so we have to obtain one fragment, we have two stripes, the digestion of Bba_K1155000 by SpeI wasn't good. | ||
+ | Well 2, 3, 4, 6, 7 : we obtain NarG, NarK and NirB in PSB1C3 and Pfnr, NarK fragments at the right size, we can purify it. | ||
+ | Well 8, 9 : we have done two digestions so we have to obtain two fragments, we have only one stripe, the digestion of Bba_K1155005 and Bba_K1155004 by EcoRI/SpeI wasn't good. | ||
+ | |} | ||
====5 - Gel purification of the digestion of Bba_K1155000 and Bba_K1155006 by EcoRI/SpeI and Bba_K1155004, Bba_K1155005 and Bba_K1155006 by SpeI==== | ====5 - Gel purification of the digestion of Bba_K1155000 and Bba_K1155006 by EcoRI/SpeI and Bba_K1155004, Bba_K1155005 and Bba_K1155006 by SpeI==== |
Revision as of 18:30, 10 September 2013
Notebook : August 14
Lab work
A - Aerobic/Anaerobic regulation system
Objective : obtaining ...
1 - Digestion of Bba_K1155004,Bba_K1155005, Bba_K1155006 by EcoRI and SpeI
- Buffer FD : 2µL
- H2O : 6µL
- DNA : 10µL
- SpeI FD : 1µL
- EcoRI FD : 1µL
2 - Electrophoresis to check the digestion of Bba_K1155004,Bba_K1155005, Bba_K1155006 by EcoRI and SpeI
Anaïs, Nadia
Expected sizes :
- PSB1C3 : 2070kb
- NarK, NarG, NirB : 200kb
We obtained NarK, NarG, NirB fragments at the right size but in very few quantity. We do it again but this time we will use more quantity of enzymes AND ADN ?!?????????? |
3 - Digestion of Bba_K1155000, Bba_K1155004,Bba_K1155005, Bba_K1155006 by SpeI and EcoRI/SpeI
Anaïs, Nadia
- Digestion by SpeI :
- Buffer : 2µL
- SpeI : 2µL
- ADN : 15µL
- H20 : 1µL
- Digestion by EcoRI and SpeI :
- Buffer : 3µL
- SpeI : 2µL
- EcoRI : 2µL
- ADN : 20µL
- H20 : 3µL
4 - Electrophoresis to check the digestion of Bba_K1155000, Bba_K1155004,Bba_K1155005, Bba_K1155006 by SpeI and EcoRI/SpeI
Anaïs, Nadia
Expected sizes :
- Pfnr in PSB1C3 : ...
- Nar K in PSB1C3, NarG in PSB1C3, NirB in PSB1C3 : ...
- PSB1C3 : 2070kb
- NarK, NarG, NirB : 200kb
- Pfnr : ...
Well 1 : we have done one digestion so we have to obtain one fragment, we have two stripes, the digestion of Bba_K1155000 by SpeI wasn't good. Well 2, 3, 4, 6, 7 : we obtain NarG, NarK and NirB in PSB1C3 and Pfnr, NarK fragments at the right size, we can purify it. Well 8, 9 : we have done two digestions so we have to obtain two fragments, we have only one stripe, the digestion of Bba_K1155005 and Bba_K1155004 by EcoRI/SpeI wasn't good. |
5 - Gel purification of the digestion of Bba_K1155000 and Bba_K1155006 by EcoRI/SpeI and Bba_K1155004, Bba_K1155005 and Bba_K1155006 by SpeI
Anaïs, Nadia
Protocol : Gel purification
Nanodrop :
- Pfnr : 13.7ng/µL
- NarK : 27.9ng/µL
- NarK in PSB1C3 : 87.1ng/µL
- NarG in PSB1C3 : 39.2ng/µL
- NirB in PSB1C3 : 37.7ng/µL
CONCLUSION
6 - Electrophoresis of the digestion of Bba_K11550007 and Bba_K1155003 by XBaI and PstI
Nadia
A - Aerobic/Anaerobic regulation system / B - PCB sensor system