Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Joan Cortada Garcia
Date created: 2014-09-12 11:00:00
Date modified: 2015-05-08 01:09:50
ybiJ promoter coupled to mKate2 reporter gene and Rhamnose promoter coupled to N-genes
| Types | DnaRegion |
| Roles | Measurement engineered_region |
| Sequences | BBa_K1316008_sequence (Version 1) |
Description
Promoter of the ybiJ gene, which is activated by the presence of several nitrogen-based compounds such as 2,4,6-TNT, 2,4-DNT and 1,3-DNB.mKate2 is a far-red fluorescent protein used with reporting purposes. mKate2 codon usage is optimised for high expression in mammalian cells (humanised). It is, nevertheless, suitable for propagation in E. coli.
nfsA, nfsB and nemA (refered here as N-genes) are the genes for NfsA, NfsB and NEM reductases, which presumably play a role in 2,4-DNT and 2,4,6-TNT metabolism. These genes are regulated by the inducible Rhamnose promoter
This construct is, then, designed to be able to detect and quantify the presence of compunds such as 2,4,6-TNT, 2,4-DNT and 1,3-DNB. The presence of the N-genes aims to enhance this response.
Notes
The Rhamnose promoter was used to inducibly regulate the expression of the N-genesA double transcriptional terminator was placed between mKate2 and the N-genes in orther to avoid unfinished transcripts of the N-genes, as well as to avoid expression of these genes without the presence of rhamnose
A strong RBS was placed in front of each gene.
An "all frame STOP" region was placed after each N-gene.
nfsA and nfsB have a 6xHis tag at the N-terminus; nemA has a 6xHis tag at the C-terminus. This way, the proteins can be purified to check if they are successfully expressed
Source
Assembly of a part from Belkin laboratory (Belkin et al. Appl Microbiol Biotechnol (2014) 98:885???895) with a "de novo" synthesised part| Access | Instance | Definition |
| public public | BBa_K1316005 BBa_K1316006 | BBa_K1316005 BBa_K1316006 |
| Sequence Annotation | Location | Component / Role(s) |
| BBa_K1316005 BBa_K1316006 | 1,1252 1261,4055 | BBa_K1316005 BBa_K1316006 |