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Team:Paris Saclay/Notebook/August/19
From 2013.igem.org
(Difference between revisions)
(→2 - Ligation of NarK, NarG or NirB with RBS-LacZ-Term or RBS-AmilCP-Term in PSB1C3) |
(→1 - Electrophoresis gel of PCR products : BphR2 Part I) |
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==='''A - Aerobic/Anaerobic regulation system'''=== | ==='''A - Aerobic/Anaerobic regulation system'''=== | ||
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| + | ===='''Objective : obtaining BBa_K1155007'''==== | ||
| + | |||
| + | ====1 - Sequence analysis of BBa_K1155007==== | ||
| + | |||
| + | N'Guyen | ||
| + | |||
| + | {| | ||
| + | | style="border:1px solid black;padding:5px;background-color:#DEDEDE;" | | ||
| + | Our sequence analysis was good. We obtain our biobrick BBa_K1155007. | ||
| + | |} | ||
===='''Objective : characterize BBa_K1155000, BBa_K1155004, BBa_K155005, BBa_K1155006 '''==== | ===='''Objective : characterize BBa_K1155000, BBa_K1155004, BBa_K155005, BBa_K1155006 '''==== | ||
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** Ligase : 1µL | ** Ligase : 1µL | ||
| - | ====3 - Transformation of NarK, NarG or NirB with RBS-LacZ-Term or RBS-AmilCP-Term in | + | ====3 - Transformation of NarK, NarG or NirB with RBS-LacZ-Term or RBS-AmilCP-Term in pSB1C3 in DH5α==== |
XiaoJing | XiaoJing | ||
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Protocol : [[Team:Paris_Saclay/Protocols/Transformation|Bacterial transformation]] | Protocol : [[Team:Paris_Saclay/Protocols/Transformation|Bacterial transformation]] | ||
| - | ====4- Transformation of MG1655Z1 Δfnr::Km==== | + | ====4- Make competent cells of strain MG1655Z1 Δfnr::Km==== |
| + | |||
| + | XiaoJing | ||
| + | |||
| + | Protocol : [[Team:Paris_Saclay/preparation|Preparation of super competent cells]] | ||
| + | |||
| + | ====5- Transformation of MG1655Z1 Δfnr::Km==== | ||
| + | |||
| + | XiaoJing | ||
| - | + | We transformed pcp20 plasmid into strain MG1655Z1 Δfnr::Km. Then we spread in LB and ampicillin antibiotic plate. We need to incubate it at 30°C for two days. | |
Protocol : [[Team:Paris_Saclay/Protocols/Transformation|Bacterial transformation]] | Protocol : [[Team:Paris_Saclay/Protocols/Transformation|Bacterial transformation]] | ||
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*Well 1 : 6µL DNA Ladder | *Well 1 : 6µL DNA Ladder | ||
| - | *Well 2 : 40µL BphR2 part I+8µl of 6X loading dye | + | *Well 2 : 40µL BphR2 part I + 8µl of 6X loading dye |
*Gel : 1% | *Gel : 1% | ||
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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 | + | We obtained a frangment at the right size. We can purify it. |
|} | |} | ||
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[[File:PSevry2.jpg]] | [[File:PSevry2.jpg]] | ||
| - | We spent our afternoon with Evry team. Each team presented its lab, modeling and human practices project. We learned informations to improve our human practices | + | We spent our afternoon with Evry team. Each team presented its lab, modeling and human practices project. We learned informations to improve our human practices project. We discussed about safety and gave them some advices about the pills they would make. |
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Latest revision as of 01:29, 5 October 2013
Notebook : August 19
Lab work
A - Aerobic/Anaerobic regulation system
Objective : obtaining BBa_K1155007
1 - Sequence analysis of BBa_K1155007
N'Guyen
|
Our sequence analysis was good. We obtain our biobrick BBa_K1155007. |
Objective : characterize BBa_K1155000, BBa_K1155004, BBa_K155005, BBa_K1155006
1 - EtOH precipitation of RBS-LacZ-Term and RBS-AmilCP-Term
Nadia
Protocol : EtOH precipitation
We used 20µL of DNA.
Nanodrop :
- RBS-LacZ-Term : 38.4ng/µL
- RBS-AmilCP-Term : 28.7ng/µL
|
The precipitation was good. We will make ligations. |
2 - Ligation of NarK, NarG or NirB with RBS-LacZ-Term or RBS-AmilCP-Term in pSB1C3
XiaoJing
Used quantities :
- NirB and RBS-LacZ-Term :
- NirB : 3µL
- RBS-LacZ-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
- NirB and RBS-AmilCP-Term :
- NirB : 3µL
- RBS-AmilCP-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
- NarG and RBS-LacZ-Term :
- NarG : 3µL
- RBS-LacZ-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
- NarG and RBS-AmilCP-Term :
- NarG : 3µL
- RBS-AmilCP-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
- NarK and RBS-LacZ-Term :
- Nar K : 3µL
- RBS-LacZ-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
- NarK and RBS-AmilCP-Term :
- NarK : 3µL
- RBS-AmilCP-Term : 1µL
- Buffer : 2µL
- H2O : 13µL
- Ligase : 1µL
3 - Transformation of NarK, NarG or NirB with RBS-LacZ-Term or RBS-AmilCP-Term in pSB1C3 in DH5α
XiaoJing
Protocol : Bacterial transformation
4- Make competent cells of strain MG1655Z1 Δfnr::Km
XiaoJing
Protocol : Preparation of super competent cells
5- Transformation of MG1655Z1 Δfnr::Km
XiaoJing
We transformed pcp20 plasmid into strain MG1655Z1 Δfnr::Km. Then we spread in LB and ampicillin antibiotic plate. We need to incubate it at 30°C for two days.
Protocol : Bacterial transformation
A - Aerobic/Anaerobic regulation system / B - PCB sensor system
Objective : obtaining FNR and BphR2 proteins
1 - Electrophoresis gel of PCR products : BphR2 Part I
Damir
|
|
Expected size :
- BphR2 Part I : 178 kb
|
We obtained a frangment at the right size. We can purify it. |
2 - Gel purification of PCR products : BphR2 Part I
Damir
Protocol : [http://www.mn-net.com/tabid/1452/default.aspx Gel purification ]
Nanodrop :
- BphR2 Part I: 57.2ng/µL
|
We lost our fragment BphR2 Part I. We will make the PCR again. |
Lab work, Modeling, Human practices
Meeting with Evry
We spent our afternoon with Evry team. Each team presented its lab, modeling and human practices project. We learned informations to improve our human practices project. We discussed about safety and gave them some advices about the pills they would make.
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