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Team:Paris Saclay/Notebook/September/2
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==Labwork== | ==Labwork== | ||
| - | === | + | ===Contruction of the fusion protein=== |
'' By Hv'' | '' By Hv'' | ||
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We first did a liquid culture in 5 mL of LB. | We first did a liquid culture in 5 mL of LB. | ||
| - | === | + | ===Lemon scent=== |
''By Melanie'' | ''By Melanie'' | ||
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Migration of the PCR done [https://2014.igem.org/Team:Paris_Saclay/Notebook/September/1#Limone_synthase yesterday] | Migration of the PCR done [https://2014.igem.org/Team:Paris_Saclay/Notebook/September/1#Limone_synthase yesterday] | ||
| - | [[File:PCR pPS5 CAD + PCR LSpGMETe 0209.jpg|500px]] | + | [[File:PCR pPS5 CAD + PCR LSpGMETe 0209.jpg|500px|center]] |
| - | [[File:PCR clone LS pGMETe (12-25) 0209.jpg|500px]] | + | [[File:PCR clone LS pGMETe (12-25) 0209.jpg|500px|center]] |
picture 1 | picture 1 | ||
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Plasmid extraction using the plasmid DNA extraction kit and electrophoresis: | Plasmid extraction using the plasmid DNA extraction kit and electrophoresis: | ||
| - | [[File:Extraction, pPS3 pPS4 0209.jpg|500px]] | + | [[File:Extraction, pPS3 pPS4 0209.jpg|500px|center]] |
We conclude that we have done a good manipulation. | We conclude that we have done a good manipulation. | ||
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====pPS5==== | ====pPS5==== | ||
To have more plasmid We do some bacteria culture (pPS5) | To have more plasmid We do some bacteria culture (pPS5) | ||
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| + | ==Photo of the Day== | ||
| + | [[File:Paris Saclay 16_september.jpg|600px|center]] | ||
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| + | {{Team:Paris_Saclay/notebook_footer}} | ||
Latest revision as of 12:36, 14 October 2014
Contents |
Tuesday 2nd September
Labwork
Contruction of the fusion protein
By Hv
We can see that the blue colonies of E.coli are not blue in a simple petri dish with ampicillin. It's a fail. So, to figure our problem we decided to do a sequencing of our plasmids.
We chose 6 colonies from an Xgal-IPTG petri dish:
- 1 completely blue
- 2 blue with a white ring around the blue
- 3 white colony
PS: we found that there is 2 types of blue colonies: completely blue and blue with a white ring around the blue)
We first did a liquid culture in 5 mL of LB.
Lemon scent
By Melanie
Limonene synthase
Migration of the PCR done yesterday
_0209.jpg)
picture 1 well 1= ladder well 2-6 = pPS5 other well = LS PCR
picture 2 = LS PCR As we see, there are nothing on our gel so we conclude that we failed to clone LS in pGMETeasy
pPPS5
the photo in the same as previously: We saw a band but we will check another time by PCR to be sure.
pPS3 and pPS4
Plasmid extraction using the plasmid DNA extraction kit and electrophoresis:
We conclude that we have done a good manipulation.
Limonene synthase PCR
As we saw that we don't have anything for LS, We try to do other PCR from the Biobrick (BBA762100). We use two enzyme: Dream taq and Vent taq
| component | volume |
|---|---|
| H2O | 42μl |
| buffer | 5μl |
| dNTPs | 1μl |
| iPS66 | 1μl |
| iPS67 | 1μl |
| DNA | 0.5μl |
| enzyme | 0.5μl |
Tubes were placed in PCR machine with the following parameters.
| Cycle step | Temperature | Time | Cycle |
|---|---|---|---|
|
Initial denaturation |
94°C |
1 min |
1 |
| Denaturation | 94°C | 30 s | 25 |
| Annealing | 50°C | 25 s | 25 |
| Extension | 72°C | 1 min | 25 |
| Final extension | 72°C | 10 min | 1 |
| Final extension | 8°C | hold | 1 |
pPS5
To have more plasmid We do some bacteria culture (pPS5)
Photo of the Day
