Team:Paris/Production testing

From 2009.igem.org

(Difference between revisions)
(→Part I : Testing delay phase I)
(→Part II : Testing delay phase II)
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|bgcolor="#F8F8F8"|'''PCR :'''  
|bgcolor="#F8F8F8"|'''PCR :'''  
-
mRFP Cterm  matrix : pSB1A3 Oligo :O59 and O60  TM :
+
terminator: igem plate
 +
pLac: igem plate
-
mRFP Nterm  plasmid : pSB1A3 Oligo :O57 and O58 TM :
 
-
|bgcolor="#E0E3FE"|'''PCR :'''
+
TetR
-
|bgcolor="#CBD1FD"|'''PCR :'''
+
 
-
|bgcolor="#BCCDFD"|'''PCR :'''
+
 
 +
pTet:
 +
 
 +
 
 +
mRFP
 +
 
 +
 
 +
|bgcolor="#E0E3FE"|-
 +
|bgcolor="#CBD1FD"|-
 +
|bgcolor="#BCCDFD"|-
|- style="background: #d8d8d8; text-align: center;"
|- style="background: #d8d8d8; text-align: center;"
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ok
ok
-
|bgcolor="#E0E3FE"|'''Verification on gel :'''
+
|bgcolor="#E0E3FE"|-
-
-
-
|bgcolor="#CBD1FD"|'''Verification on gel :'''
+
|bgcolor="#CBD1FD"|-
-
-
-
|bgcolor="#BCCDFD"|'''Verification on gel :'''
+
|bgcolor="#BCCDFD"|-
-
-
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ok
ok
-
|bgcolor="#E0E3FE"|'''Purification on gel :'''
+
|bgcolor="#E0E3FE"|
-
-
-
|bgcolor="#CBD1FD"|'''Purification on gel :'''
+
|bgcolor="#CBD1FD"|
-
-
-
|bgcolor="#BCCDFD"|'''Purification on gel :'''
+
|bgcolor="#BCCDFD"|
-
-
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|bgcolor="#F8F8F8"|'''Digestion:'''  
|bgcolor="#F8F8F8"|'''Digestion:'''  
-
TatABCE
+
pLac  S/P
 +
 
 +
 
 +
TetR X/P
 +
 
 +
 
 +
terminator  E/X
 +
 
 +
 
 +
pTet  S/P
 +
 
 +
 
 +
mRFP  X/P
 +
 
 +
 
 +
pSB3T5  E/P
|bgcolor="#E0E3FE"|'''Digestion:'''  
|bgcolor="#E0E3FE"|'''Digestion:'''  
 +
plac-TetR  E/S
 +
 +
pTet-mRFP  X/P
 +
|bgcolor="#CBD1FD"|'''Digestion:'''  
|bgcolor="#CBD1FD"|'''Digestion:'''  
 +
pLac-tetR-ter  S/P
 +
 +
|bgcolor="#BCCDFD"|'''Digestion:'''  
|bgcolor="#BCCDFD"|'''Digestion:'''  
 +
pLac-tetR-ter-pTet-mRFP  E/P
 +
|- style="background: #bebebe; text-align: center;"   
|- style="background: #bebebe; text-align: center;"   

Revision as of 01:29, 22 October 2009

iGEM > Paris > WetLab > Production


Contents

WetLab - Production

  1. The delay
    1. Part I : Testing delay phase I
    2. Part I : Functional testing of the part I
    3. Part II : Testing delay phase II
    4. Part II : Functional testing of the part II
  2. The destabilization of the membrane

The delay

We separate the construction on two plasmids : Part I and Part II.

Part I : Testing delay phase I

The green color means the experiment was a success


plasmid = PSB3T5



Experiments ran :


Column 1 Column 2 Column 3 Column 4
PCR :

pBAD: plate igem


LacI: plate igem


Terminator: plate igem


RBS: plate igem


YFP: plate igem

- - -
Verification on gel :

ok

- - -
Purification on gel :

ok

- - -



Digestion:

pBAD S/P


lacI E/S


terminator E/X


RBS S/P


YFP X/P

Digestion:

pBAD-LacI E/S


RBS-YFP E/S

Digestion:

RBS-YFP-ter E/X

-
Verification digestion:

ok

Verification digestion:

ok

Verification digestion:

ok

-


Ligation:

pBAD-LacI

RBS-YFP

Ligation: Ligation: Ligation:
Colony PCR :

ok

Colony PCR :

ok

Colony PCR :

ok

Colony PCR:

ok

Miniprep:

clone

Miniprep:

clone

Miniprep:

clone

Miniprep:

clone

Sequencing :

ok

Sequencing :

ok

Sequencing :

-

Sequencing :

-

Stock glycerol:


Stock glycerol

Cly A(Cter)-(Nter)RFP: S72(clone 8)

Stock glycerol

-

Stock glycerol

-

Part I : Functional testing of the part I

PBAD LacI LVA- YFP was transforme into Top ten Bacteria and grew on medium :


1% arabinose

1% glucose

LB pure


Part II : Testing delay phase II

The green color means the experiment was a success


plasmid =



Experiments ran :


Column 1 Column 2 Column 3 Column 4
PCR :

terminator: igem plate

pLac: igem plate


TetR


pTet:


mRFP


- - -
Verification on gel :

ok

-

-

-

-

-

-

Purification on gel :

ok

-

-

-


Digestion:

pLac S/P


TetR X/P


terminator E/X


pTet S/P


mRFP X/P


pSB3T5 E/P

Digestion:

plac-TetR E/S

pTet-mRFP X/P

Digestion:

pLac-tetR-ter S/P


Digestion:

pLac-tetR-ter-pTet-mRFP E/P


Verification digestion:

ok

Verification digestion:

-

Verification digestion:

-

Verification digestion:

-


Ligation:

TatABCE

Ligation: Ligation: Ligation:
Colony PCR:

ok

Colony PCR :

-

Colony PCR:

-

Colony PCR :

-

Miniprep:

STOPPED

Miniprep:

-

Miniprep:

-

Miniprep:

-

Sequencing :

-

Sequencing :

-

Sequencing :

-

Sequencing :

-

Stock glycerol:

-

Stock glycerol

-

Stock glycerol

-

Stock glycerol

-

Part II : Functional testing of the part II

Ptet RFP Plac TetR was tranforme into Top10 bacteria and grew on media :


0,1% IPTG .

1% glucose.

LB pure.



The destabilization of the membrane

The green color means the experiment was a success


plasmid = PSB3T5



Experiments ran :


Column 1 Column 2 Column 3 Column 4
PCR :

TatABCE matrix :

PCR : PCR : PCR :
Verification on gel :

ok

Verification on gel :

-

Verification on gel :

-

Verification on gel :

-

Purification on gel :

ok

Purification on gel :

-

Purification on gel :

-

Purification on gel :

-


Digestion:

TatABCE

Digestion: Digestion: Digestion:
Verification digestion:

ok

Verification digestion:

-

Verification digestion:

-

Verification digestion:

-


Ligation:

TatABCE

Ligation: Ligation: Ligation:
Colony PCR :

ok

Colony PCR:

-

Colony PCR:

-

Colony PCR:

-

Miniprep:

STOPPED

Miniprep:

-

Miniprep:

-

Miniprep:

-

Sequencing :

-

Sequencing :

-

Sequencing :

-

Sequencing :

-

Stock glycerol:

-

Stock glycerol

-

Stock glycerol

-

Stock glycerol

-



Retrieved from "http://2009.igem.org/Team:Paris/Production_testing"