Difference between revisions of "Team:UPMC-Paris/Part Characterization"

BBa_K2180001 : Zinc Operator - CzrA Repressor Binding Site

http://parts.igem.org/Part:BBa_K2180001
Biobrick based on a previous biobrick made by the iGEM09_Newcastle Team (2009-10-14) :

http://parts.igem.org/Part:BBa_K174015

We add two restriction sites: SacII before and PacI after the regulatory region in order to use it in our project. They allow us to replace a regulatory region with another regulatory region.

 

BBa_K2180002 : Fructose Operator - FruR Binding Site

http://parts.igem.org/Part:BBa_K2180002
The sequence of the regulatory region comes from this site : http://regprecise.lbl.gov/RegPrecise/regulon.jsp?regulon_id=12681
We add two restriction sites: SacII before and PacI after the regulatory region in order to use it in our project. They allow us to replace a regulatory region with another regulatory region. The regulatory region is composed by two FruR binding sites wich are derepressed in the presence of fructose.

 

BBa_K2180003 : Sucrose Operator - DegU Binding Site

http://parts.igem.org/Part:BBa_K2180003
The sequence of the regulatory region comes from this site : http://www.prodoric.de/site.php?site_acc=SI002871
We add two restriction sites SacII before and PacI after the regulatory region in order to use it in our project. They allow us to replace a regulatory region with another regulatory region. The regulatory region is composed by two DegU binding (one forward, one reverse) sites wich are derepressed in the presence of sucrose.

 

BBa_K2180004 : LacI + LVA for B. subtilis

http://parts.igem.org/Part:BBa_K2180004

The sequence is based on a previous IGEM part made by the group Antiquity (2003-01-31) coding for LacI + LVA for E. coli:

http://parts.igem.org/Part:BBa_C0012

Coding sequence of LacI protein + LVA (rapid degradation tail) wich improve the switch time and make it more sensitive.

We had to improve the codon usage for B. subtilis

 

BBa_K2180005 : Blue Pigment for B. subtilis

http://parts.igem.org/Part:BBa_K2180005

Biobrick based on a previous biobrick made by the Team iGEM11_Uppsala-Sweden   (2011-09-18) :

http://parts.igem.org/wiki/index.php?title=Part:BBa_K592009

This sequence is coding for a blue pigment we use in our detection system as reporter.

We had to improve the codon usage for B. subtilis.

 

BBa_K2180006 : Promoter LacO for B. subtilis

http://parts.igem.org/Part:BBa_K2180006

The part is made of a previous IGEM part made by the team iGEM12_LMU-Munich   (2012-07-16) :

http://parts.igem.org/Part:BBa_K823002

We add two LacI binding sites (Gilbert and Maxam, PNAS, 1973)

This part is made of PlepA promoter, two LacI binding sites and a RBS. There is a PacI restriction site before LacI binding sites and a SacII restriction site after them. They allows us to remplace the LacI binding sites by another operating sequence. A SalI restriction sites is presents before the PlepA sequence and a NdeI restriction site after the RBS that may be used to insert the whole sequence. We use this part in our project to promote the pigment expression under the control of LacI.

 

BBa_K2180007 : Promoter Fur for B. subtilis

http://parts.igem.org/Part:BBa_K2180007

This part is based on a previous IGEM part made by the Team iGEM12_LMU-Munich  (2012-07-16) :
http://parts.igem.org/wiki/index.php?title=Part:BBa_K823000
We add the Fur Box site (Baichoo and Helmann, J. Bacteriol., 2002) in order to make this promoter inactive in the absence of Iron.

This part is made of PliaG promoter, a fur (Ferric Uptake Regulator) binding site and a RBS. There is a ClaI restriction site before the Fur Box (Fur Binding Site) and a PscI restriction site after. They allows us to remplace Fur Box by another operating sequence. A BamHI restriction sites is present before the PliaG sequence and a NcoI restriction site after the RBS that may be used to insert the whole sequence. We use this part in our project to promote the LacI + LVA expression under the control of Fur.
We had to check the compatibility of the promoter with the Fur Box.

 

BBa_K2180008 : Double terminator

http://parts.igem.org/Part:BBa_K2180008

This part is based on a previous iGEM part made by the Team Antiquity (2003-07-17) :

http://parts.igem.org/wiki/index.php?title=Part:BBa_B0015
This is a double terminator we use in our project for the pigment production. He is made of a transcriptional terminator in forward and reverse direction. We add a BmtI restriction site before the first terminator (forward) and an AvrII restriction site after the second terminator (reverse).

 

BBa_K2180009 : LacI + LVA + Promoter + Terminator

http://parts.igem.org/Part:BBa_K2180009
This is the assembly of our basic parts BBa_K2180004, BBa_K2180007 and BBa_K2180008

 

BBa_K2180010 : Cadmium Operator - MntR Binding Site

http://parts.igem.org/Part:BBa_K2180010

Biobrick based on a previous biobrick made by the iGEM09_Cornell Team (2009-08-10) :

http://partsregistry.org/Part:BBa_K285103:Design

We add two restriction sites: SacII before and PacI after the regulatory region in order to use it in our project. They allow us to replace a regulatory region with another regulatory region. This Region allow MntR to bind to DNA and repress transcription. In the presence of Cadmium transcription is derepressed because of the fixation of cadmium to MntR.

*Other metallic ions may bind to MntR.

 

BBa_K2180011 : Cadmium Operator - ArsR Binding Site

http://parts.igem.org/Part:BBa_K2180011

Biobrick based on a previous biobrick made by the iGEM09_Newcastle Team (2009-10-14) :

http://parts.igem.org/Part:BBa_K174015

We add two restriction sites: SacII before and PacI after the regulatory region in order to use it in our project. They allow us to replace a regulatory region with another regulatory region. This Region allows ArsR to bind to DNA and repress transcription. In the presence of Heavy Metals such as Arsenic or Cadmium transcription is derepressed because of the fixation to ArsR.

*Other metalic ions may bind to ArsR.

 

BBa_K2180012 : Blue Pigment+ PLacO + Double terminator

http://parts.igem.org/Part:BBa_K21800102

This is the assembly of our parts BBa_K2180005, BBa_K2180006 and BBa_K2180008