Part:BBa_K1909014
Omp Promoter / eYFP
The part BBa_K1909014 consists of the yellow fluorescent Protein eYFP and the positively regulated OmpR-controlled promoter (BBa_R0082). The promoter OmpR is taken from the upstream region of ompC. Phosphorylated OmpR binds to the three operator sites and activates transcription.
Usage and Biology
In nature, the promoter OmpR is upstream of the ompC porin gene. The regulation of ompC is determined by the EnvZ-OmpR osmosensing machinery. EnvZ phosphorylates OmpR to OmpR-P. At high osmolarity, EnvZ is more active, creating more OmpR-P. OmpR-P then binds to the low-affinity OmpR operator sites upstream of ompC. Via FACS analysis it was shown in EnvZ deficient cells (JW3367-3), containing the construct BBa_K1909014, that the expression of eYFP was significantly reduced in comparison to EnvZ containing cells (DH5α).
Functional Parameters
In two approaches, BBa_R0082 (OmpR, positive promoter) was ligated with BBa_E0430 (eYFP) into pSB1C3 and transformed the constructs into both ΔEnvZ (JW3367-3) and wt (DH5a) E. coli strains. Positive constructs were validated by sequencing. Negative controls (JW3367-3) and DH5a without our constructs) showed only few to no fluorescence as expected (fig.1 A, B). EYFP expression (fig. 2 A) proved to be significantly higher in DH5a (green) cells as compared to EnvZ deficient JW3367-3 cells (pink).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Functional Parameters
| chassis | E. coli JW 3367-3 |
//cds/reporter/gfp
| chassis | E. coli JW 3367-3 |