Team:Paris Saclay/Notebook/September/8

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{{Team:Paris_Saclay/notebook_header}}
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=Monday 8th September=
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==Lab Work==
==Lab Work==
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''by Mélanie''
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===D-Lemon Scent===
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===Lemon Scent===
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====Checking PCR====
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Electrophoresis of the PCR made [http://2014.igem.org/Team:Paris_Saclay/Notebook/September/5#PCR_LS_PS_GS_Cad Friday]
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[photo]
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====Cloning====
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With this PCR, I do a cloning in a Topo Cloning vector (Zero Blunt TOPO PCR Cloning Kit) (LS GS and CAD)
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{| class="wikitable centre" width="50%"
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|+
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|-
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! scope=col | component
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! scope=col | volume
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|-
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|PCR product
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|1μl
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|-
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|Salt
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|1μl
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|-
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|Vector
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|1 μl
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|-
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|H<sub>2</sub>O
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|3μl
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|}
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Between 5 and 30 min at room temperature
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==== Transformation====
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Add 50µL of bacteria to the clonage previously done
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30min on ice
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45 sec at 42°C
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add 1ml of LB
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1hour at 37°
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Spread on a petri dish
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====PCR with bacteria====
====PCR with bacteria====
From the petri dish ([http://2014.igem.org/Team:Paris_Saclay/Notebook/September/5#Transformation made here], I select some clone to do a PCR :
From the petri dish ([http://2014.igem.org/Team:Paris_Saclay/Notebook/September/5#Transformation made here], I select some clone to do a PCR :
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[photo]
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[[File:0809 PCR clones.jpg|400px|center]]
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We identify that we have success with to clone of PS.
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We identify that we have success with to clone of PS. (well 5-6)
====Liquide culture of clones====
====Liquide culture of clones====
The PS clone that reveal to be ok are cultivate in LB + AMP médium
The PS clone that reveal to be ok are cultivate in LB + AMP médium
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===B-Contruction of the fusion protein===
+
====Checking PCR====
 +
Electrophoresis of the PCR made [http://2014.igem.org/Team:Paris_Saclay/Notebook/September/5#PCR_LS_PS_GS_Cad Friday]
 +
 
 +
LS PS GS CAD (and chromo) (2 enzymes = vent taq and dream taq)
 +
 
 +
[[File:0809 PCR LSPSGSCADCRHOMO vent dream.jpg|400px|center]]
 +
 
 +
====Cloning====
 +
With this PCR, I do a cloning in a Topo Cloning vector (Zero Blunt TOPO PCR Cloning Kit) (LS GS and CAD)
 +
 
 +
{| class="wikitable centre" width="50%"
 +
|+
 +
|-
 +
! scope=col | component
 +
! scope=col | volume
 +
|-
 +
|PCR product
 +
|1μl
 +
|-
 +
|Salt
 +
|1μl
 +
|-
 +
|Vector
 +
|1 μl
 +
|-
 +
|H<sub>2</sub>O
 +
|3μl
 +
|}
 +
 
 +
 
 +
Between 5 and 30 min at room temperature
 +
 
 +
==== Transformation====
 +
 
 +
Add 50µL of bacteria to the clonage previously done
 +
 
 +
30min on ice
 +
45 sec at 42°C
 +
 
 +
add 1ml of LB
 +
 
 +
1hour at 37°
 +
 
 +
Spread on a petri dish
 +
 
 +
===Construction of the fusion protein===
We check the sequence of the fusion protein and identify a clone with our chromoprotein = W6
We check the sequence of the fusion protein and identify a clone with our chromoprotein = W6
 +
 +
==Photo of the Day==
 +
[[File:Paris Saclay 8_september.jpg|150px|center]]
 +
 +
[http://ifris.org/membre/morgan-meyer/ Morgan Meyer] , one of those few we have interviewed during this month of september.
 +
 +
To learn more about the ethic aspect of our project, just click [http://2014.igem.org/Team:Paris_Saclay/Ethics here]
 +
 +
 +
{{Team:Paris_Saclay/notebook_footer}}

Latest revision as of 12:55, 14 October 2014

Contents

Monday 8th September

Lab Work

by Mélanie

Lemon Scent

PCR with bacteria

From the petri dish (made here, I select some clone to do a PCR : clones 1-2-3 for cad clones 4-5 for PS clones 6 for LS

component volume
H2O 41.5μl
buffer 5μl
dNTPs 1μl
Primer 1 1μl
Primer 2 1μl
bacteria (about 1µl = 1 colony)
Dream taq 0.5μl

Primer used:

For cloning in Topo vector = Pu Pr (universal primer)

For pPS2 = iPS 66/67

Cycle step Temperature Time Cycle

Bacteria lysis

95°C

5 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
0809 PCR clones.jpg

We identify that we have success with to clone of PS. (well 5-6)

Liquide culture of clones

The PS clone that reveal to be ok are cultivate in LB + AMP médium

Checking PCR

Electrophoresis of the PCR made Friday

LS PS GS CAD (and chromo) (2 enzymes = vent taq and dream taq)

0809 PCR LSPSGSCADCRHOMO vent dream.jpg

Cloning

With this PCR, I do a cloning in a Topo Cloning vector (Zero Blunt TOPO PCR Cloning Kit) (LS GS and CAD)

component volume
PCR product 1μl
Salt 1μl
Vector 1 μl
H2O 3μl


Between 5 and 30 min at room temperature

Transformation

Add 50µL of bacteria to the clonage previously done

30min on ice 45 sec at 42°C

add 1ml of LB

1hour at 37°

Spread on a petri dish

Construction of the fusion protein

We check the sequence of the fusion protein and identify a clone with our chromoprotein = W6

Photo of the Day

Paris Saclay 8 september.jpg

Morgan Meyer , one of those few we have interviewed during this month of september.

To learn more about the ethic aspect of our project, just click here


Back to the calendar