Part:BBa_K2278011
The alpha acetolactate synthase (ALS) enzyme catalyzes the conversion of pyruvate to acetolactate, which is then spontaneously oxidized as diacetyl. This enzyme is crucial in the synthesis pathway of diacetyl in Lactococcus lactic.
As diacetyl is not produced by wild type Vibrio harveyi, the gene responsible for ALS production is inserted in a genetic construction downstream the pTet promoter.
The pTet repressible promoter enables a compatibility with the TetR/pTet inverter system and a constitutive transcription of the gene in absence of tetR.
In the INSA UPS France 2017 project, this part is used to establish an eukaryote-prokaryote communication system based on diacetyl so that Vibrio harveyi could trigger an engineered production pathway of the Yeast Pichia pastoris.
pTet driven Diacetyl generator (pTet + ALS)
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1738
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Introduction
This DNA biobrick was designed in order to produce in strain.
1- Biological background
Mécanisme2- Usage in iGEM projects
The BBa_K2278011 cames from the module of the Croc’n cholera project (team INSA-UPS-France 2017) It was designed to produce
The part includes
Experiments
1- Molecular biology
The gene was placed in silico under the control of the p promoter (BBa_R), a strong RBS (BBa_B0034) and a terminator (BBa_B1006). IDT performed the DNA synthesis and delivered the part as gBlock. The construct was cloned by conventional ligation into pSB1C3 plasmid and transformed into E. coli Dh5 alpha strain. X transformants were obtained.
Analysis of the restriction mapSequencing
The sequencing show a2- Expression in vivo
sous titre
Protocole
Characterization
1- Validation of
descriptionmanip1
Image styléeInterprétation
manip2
Image styléeinterprétation
Discussion :
2. 2ème approche
brillante analyse
Discussion :
des perspectives éclectiques
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