Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Joachim Steen Larsen
Date created: 2016-10-04 11:00:00
Date modified: 2016-10-05 06:43:43
Beta-Ketothiolase fused to Cyan Fluorescent Protein
| Types | DnaRegion |
| Roles | engineered_region Composite |
| Sequences | BBa_K1975004_sequence (Version 1) |
Description
This is a propionate-CoA transferase (commonly known as PhaA). This enzyme catalyzes the first step in the production of PHB. The reaction it catalyzes is two molecules of acetyl-CoA is being putting together to one molecule of acetoacetyl-CoA under the usage of a propanoate that will be converted to acetate.By combining it with the synthetic gene, LPE it can be used in the synthesis of P(LA-co-3HB). This gene is fused with the Cyan Fluorescence Protein with a small linker of 6 bp between. The fused CFP is used to visual the expression of the protein. Hereby, the job will be easier to screen for positive transformants after induction (if such is used as in our case). The sequence for PhaA is taken from the BioBrick Part:BBa_K759004 and is optimized to be expressed in Bacillus subtilis.
Notes
To secure the optimal expression we choose to make a codon optimization of the genes. This was done through the Codon Optimization tool at IDT's webpage.Besides that we looked through the literature to figure our if the PhaA and CFP should be directly attached or if they should have some kind of linker. After looking through different studies that have been working with PhaA in Escherichia coli we came to the conclusion that a linker should not be necessary to secure high expression and activity. We therefore decided to only make a small linker (6 bp) between the gene and the tag.
Source
The genetic sequence for PhaA is original from Ralstonia eutropha genomic DNA, but has been optmiized to the expression of B. subtilis.The sequence for CFP genomic DNA from Acropora aculeus. This sequence has also been optimized for expression in B. subtilis.
| Access | Instance | Definition |
| public public | BBa_K1975002 BBa_K1975005 | BBa_K1975002 BBa_K1975005 |
| Sequence Annotation | Location | Component / Role(s) |
| BBa_K1975002 BBa_K1975005 | 1,1182 1189,1893 | BBa_K1975002 BBa_K1975005 |