Difference between revisions of "Part:BBa K3771042"
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<img src="https://2021.igem.org/wiki/images/c/c9/T--NCKU_Tainan--taurine_pathway_1.png" style="width:60%;"> | <img src="https://2021.igem.org/wiki/images/c/c9/T--NCKU_Tainan--taurine_pathway_1.png" style="width:60%;"> | ||
</div> | </div> | ||
− | <p align="center">Fig.1 Taurine pathways in <i>E. coli</i></p> | + | <p align="center">Fig. 1. Taurine pathways in <i>E. coli</i></p> |
<br><b style="font-size:1.3rem">Usage</b><br> | <br><b style="font-size:1.3rem">Usage</b><br> | ||
<br>We ligased the <i>cdo1-6xHis</i> fragment and <i>pspA</i> promoter on the pSU expression vector and transformed it into DH5α to complete construction of the plasmid. | <br>We ligased the <i>cdo1-6xHis</i> fragment and <i>pspA</i> promoter on the pSU expression vector and transformed it into DH5α to complete construction of the plasmid. |
Revision as of 03:13, 22 October 2021
PpspA-CDO1-6xHis
Description
This composite part is a component of the IFN-γ sensing system and was used to express the taurine production enzyme, CDO1.
Biology
Binding of IFN-γ to the OmpA/OprF chimeric protein induces the response of the phage shock protein (Psp) system, a highly conserved stress response system in enterobacteria. [1] Signal transduction from the outer membrane to the inner membrane activates the pspA promoter, initiating expression of CDO1-his-tag. CDO1 converts L-cysteine into L-cysteine sulfonic acid in the taurine synthesis L-cysteine sulfinic acid pathway [2].
Fig. 1. Taurine pathways in E. coli
Usage
We ligased the cdo1-6xHis fragment and pspA promoter on the pSU expression vector and transformed it into DH5α to complete construction of the plasmid. The his-tag allows for confirmation of CDO1 expression by western blot using the anti-6X his-tag antibody.
Characterization
Fig. 2. Colony PCR confirmation of the construction
Reference
1. Darwin AJ. The phage-shock-protein response. Molecular Microbiology. 2005;57(3):621-628. doi:10.1111/j.1365-2958.2005.04694.xhttps://pubmed.ncbi.nlm.nih.gov/16045608/
2. Joo Y-C, Ko YJ, You SK, et al. Creating a New Pathway in Corynebacterium glutamicum for the Production of Taurine as a Food Additive. Journal of Agricultural and Food Chemistry. 2018;66(51):13454-13463. doi:10.1021/acs.jafc.8b05093https://pubmed.ncbi.nlm.nih.gov/30516051/
Sequence and Features
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 12
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 591
- 1000COMPATIBLE WITH RFC[1000]