Part:BBa_K1962012
A device for expression of Im-E3 in response to bile salts
This is a composite part that is designed to induce production of the Immunity Protein to Colicin E3 in the presence of bile. The composite part consists of the bile salt responsive promoter (BBa_K318514), RBS (BBa_B0030) and the Colicin E3 Immunity protein (BBa_K1962004). For best results, use in conjunction with the transcriptional activator RamA (BBa_K1962009).
Usage and Biology
The immunity proteins for colicin Ia and E3 were cloned in front of the acrRA bile salt promoter (BBa_K1962011 and BBa_K1962012. A HA tag was fused onto the C-terminal of the immunity proteins to allow visualisation of expression via western blotting (Fig 1).
Figure 1. Western blot of colicin Ia and E3 immunities with bile salt sensitive pacrRA: The col Ia and E3 immunities were amplified with a HA tag to detect expression via a western blot. Empty pSB1C3 and acrRA promoter was blotted alongside as a control. The constructs with ramA were also blotted to see if this has any effect on acrRA function. No expression of acrRA col Ia immunity was detected however we were able to see expression of the E3 immunity in the acrRA col E3 immunity with and without ramA.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 471
Illegal XhoI site found at 785 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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