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Team:Paris/Production design
From 2009.igem.org
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==== Biobricks ==== | ==== Biobricks ==== | ||
| - | Expression of the proteins targeted to OMVs as well as ''lacI'' is controled by the Pbad promoter [http://partsregistry.org/Part:BBa_K113009 BBa_K11309]. | + | |
| + | * Expression of the proteins targeted to OMVs as well as ''lacI'' is controled by the Pbad promoter [http://partsregistry.org/Part:BBa_K113009 BBa_K11309]. | ||
We used the Ptet promoter [http://partsregistry.org/Part:BBa_R0040 BBa_ROO40] and the coding sequence for the TetR repressor protein [http://partsregistry.org/Part:BBa_C0040 BBa_COO40] to tightly repress the expression of protein domains that are expected to destabilize the outer membrane, thereby leading to increased production of OMVs. We hope that TetR mediated repression is strong enough to avoid leakage and weak expression of these deleterious polypeptides. | We used the Ptet promoter [http://partsregistry.org/Part:BBa_R0040 BBa_ROO40] and the coding sequence for the TetR repressor protein [http://partsregistry.org/Part:BBa_C0040 BBa_COO40] to tightly repress the expression of protein domains that are expected to destabilize the outer membrane, thereby leading to increased production of OMVs. We hope that TetR mediated repression is strong enough to avoid leakage and weak expression of these deleterious polypeptides. | ||
| - | The Plac promoter [http://partsregistry.org/Part:BBa_I14032 BBa_I14032] associated to ''lacI'' [BBa_C0012 http://partsregistry.org/Part:BBa_C0012] to control the expression of ''tetR''. | + | |
| - | The double terminator [http://partsregistry.org/Part:BBa_B0014 BBa_B0014] was used to isolates the different expression units from each other. | + | * The Plac promoter [http://partsregistry.org/Part:BBa_I14032 BBa_I14032] associated to ''lacI'' [BBa_C0012 http://partsregistry.org/Part:BBa_C0012] to control the expression of ''tetR''. |
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| + | * The double terminator [http://partsregistry.org/Part:BBa_B0014 BBa_B0014] was used to isolates the different expression units from each other. | ||
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| + | ==== Our contributions ==== | ||
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| + | * clyA fusion N-term and C-term | ||
| + | * | ||
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| + | '''NB:''' All coding sequences intented to be expressed are associated with a standard RBS incorporated in the forward primer used to amplify the sequence by PCR. The standard RBS used was carefully designed to ensure maximal functionality (see xxx) | ||
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The RBS were designed by Guillaume Cambray and integrated directly inside the primers. | The RBS were designed by Guillaume Cambray and integrated directly inside the primers. | ||
Revision as of 13:07, 8 September 2009
iGEM > Paris > Production > Parts Design
Contents |
Parts
Registered parts we used
Backbones
The function "increased production of OMV" is borne on two plasmid backbones pSB1A3 and pSB3T5.
Biobricks
- Expression of the proteins targeted to OMVs as well as lacI is controled by the Pbad promoter BBa_K11309.
We used the Ptet promoter BBa_ROO40 and the coding sequence for the TetR repressor protein BBa_COO40 to tightly repress the expression of protein domains that are expected to destabilize the outer membrane, thereby leading to increased production of OMVs. We hope that TetR mediated repression is strong enough to avoid leakage and weak expression of these deleterious polypeptides.
- The Plac promoter BBa_I14032 associated to lacI [BBa_C0012 http://partsregistry.org/Part:BBa_C0012] to control the expression of tetR.
- The double terminator BBa_B0014 was used to isolates the different expression units from each other.
Our contributions
- clyA fusion N-term and C-term
NB: All coding sequences intented to be expressed are associated with a standard RBS incorporated in the forward primer used to amplify the sequence by PCR. The standard RBS used was carefully designed to ensure maximal functionality (see xxx)
