Difference between revisions of "Team:Ionis Paris/Parts"

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The biosensor is the heart of our project and because it took us a lot of time to create. Like it is already explain in the <a href="https://2016.igem.org/Team:Ionis_Paris/Biobrick_Design">Biobrick Design</a> section, this biobrick is composed of the Pr promoter (<a href="http://parts.igem.org/Part:BBa_K2023004" ><font color="DeepPink">BBa_K2023004</font></a>) which is a constitutive promoter driving the transcription of the XylR gene (<a href="http://parts.igem.org/Part:BBa_K2023005" ><font color="DeepPink">BBa_K2023005</font></a>) coding for the XylR protein. The XylR protein is able to bind aromatic hydrocarbons that carry a methyl group like toluene and xylene. When XylR has bind toluene for instance, it is able to regulate the Pu promoter has a transcriptional regulator. Then, Pu is activated and it allows the transcription of the bioluminescent reporter gene GLuc (Pu+GLuc: <a href="http://parts.igem.org/Part:BBa_K2023003" ><font color="DeepPink">BBa_K2023003</font></a>), coding for the Gaussia luciferase. When this enzyme reacts with its substrate luciferine, a substance called Coelenterazine, it emits luminescence.
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The biosensor is the heart of our project and because it took us a lot of time to create. Like it is already explain in the <a href="https://2016.igem.org/Team:Ionis_Paris/Biobrick_Design"><font color="DeepPink">Biobrick Design</font></a> section, this biobrick is composed of the Pr promoter (<a href="http://parts.igem.org/Part:BBa_K2023004" ><font color="DeepPink">BBa_K2023004</font></a>) which is a constitutive promoter driving the transcription of the XylR gene (<a href="http://parts.igem.org/Part:BBa_K2023005" ><font color="DeepPink">BBa_K2023005</font></a>) coding for the XylR protein. The XylR protein is able to bind aromatic hydrocarbons that carry a methyl group like toluene and xylene. When XylR has bind toluene for instance, it is able to regulate the Pu promoter has a transcriptional regulator. Then, Pu is activated and it allows the transcription of the bioluminescent reporter gene GLuc (Pu+GLuc: <a href="http://parts.igem.org/Part:BBa_K2023003" ><font color="DeepPink">BBa_K2023003</font></a>), coding for the Gaussia luciferase. When this enzyme reacts with its substrate luciferine, a substance called Coelenterazine, it emits luminescence.
  
 
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Revision as of 21:40, 18 October 2016

Our biosensor

Part Name Nickname Type Designer Lengh
BBa_K2023001 Biosensor device BB123 Measurement C.Chenebault, C.Soucies, B.Piot 3299 bp

The biosensor is the heart of our project and because it took us a lot of time to create. Like it is already explain in the Biobrick Design section, this biobrick is composed of the Pr promoter (BBa_K2023004) which is a constitutive promoter driving the transcription of the XylR gene (BBa_K2023005) coding for the XylR protein. The XylR protein is able to bind aromatic hydrocarbons that carry a methyl group like toluene and xylene. When XylR has bind toluene for instance, it is able to regulate the Pu promoter has a transcriptional regulator. Then, Pu is activated and it allows the transcription of the bioluminescent reporter gene GLuc (Pu+GLuc: BBa_K2023003), coding for the Gaussia luciferase. When this enzyme reacts with its substrate luciferine, a substance called Coelenterazine, it emits luminescence.


Favorite Parts

Part Name Nickname Type Designer Lengh
BBa_K2023004 Pr-RBS BB1 Composite C.Chenebault 448 bp
Bba_K2023005 XylR BB2 Coding C.Soucies 1704 bp
BBa_K2023006 Term-Pu-RBS-Gluc-term BB3 Composite B.Piot 1330 bp

Those 3 parts are probably the most important because they are the fondations of the biosensor right above. We designed and ordered the sequences to Integrated DNA Technologies. We first assembled BB1 and BB2 in pSB1C3 to create BB12 and then we had BB3 in order to produce BB123 the biosensor. For more information on those parts clic on the Biobricks to go to the registry.

Part Name Nickname Type Designer Lengh
BBa_K2023002 XylR coding device BBA Regulatory C.Chenebault 2261 bp
BBa_K2023003 Gluc coding device with Pu BBB Reporter C.Soucies 1028 bp

BBA and BBB are the coding device that compose our biosensor. BBA is made with Pr-RBS-XylR-Term and can be linked with other reporter like the GFP reporter. BBB is compesed of GLuc coding device with Pu: RBS-GLuc-Term-Pu.


Parts for CelloCad

Part Name Nickname Type Designer Lengh
Bba_K2023014 GFP coding device with Pr C2 Device V.Plait 1314
Bba_K2023015 XylR coding device-GFP coding device with Pu C3 Measurement C.Soucies 3391 bp
Bba_K2023017 Pu-RBS C4 Composite C.Chenebault 358 bp
Bba_K2023016 GFP coding device with Pu C45 Reporter C.Soucies 1224 bp

Those parts allowed us to start the CelloCad side project. In order to compare the RPU of the Pr and the Pu promoter with the reference promoter BBa_J23101 we designed C2 and C3. To produce the part C3 we have needed the parts C4 and C45.



Gaussia derived parts

Part Name Nickname Type Designer Lengh
Bba_K2023007 Gluc coding device with Pr G1 device B.Piot 1120 bp
Bba_K2023008 Gluc-term G2 Composite C.Soucies 663 bp
Bba_K2023009 Gluc codon optimized G3 Coding B.Piot 561 bp

G1, G2 and G3 are 3 parts that come from the GLuc-His part of 522 bp (BBa_K1732027). G1 will allow a positive control during a futur pollution quantification test. G2 is a useful part because it is the GLuc-His part with a terminator and it can be adapte with another detector than the XylR. G3 is the GLuc-His part optimized for our project in fonction of our chassis which is E.Coli and for the IDT sequence order.


XylR derived parts

Part Name Nickname Type Designer Lengh
Bba_K2023010 XylR with His tag X2 Coding C.Soucies 1667
Bba_K2023011 XylR-His tag coding device X3 Regulatory C.Soucies 2319 bp
Bba_K2023012 BB1- XylR-His tag BB12His Composite C.Chenebault 1864 bp
Bba_K2023013 XylR coding device with mRFP X1 Regulatory B.Piot 3023 bp

Those 4 parts are derived from the XylR gene (BB2) that we optimized for our chassis. X2 is the XylR with a His Tag. We designed this part in order to purify and quantify XylR concentration in ou medium during a test. BB12His is X2 plus the promoter Pr. The coding device is X3: it is composed of BB12His plus a terminator. In the same way of monitoring XylR, we designed XylR coding device with mRFP. This device is composed of Pr-RBS-XylR-RBS-mRFP-Terminator. It allow to prove that the XylR is synthesized in the medium when the colonies becomes red.