Part:BBa_K1962015
A device for expression of Im-Ia in response to pH
This is a composite part for the production of the Immunity Protein to Colicin Ia in response to environmental pH changes. The composite part consists of a pH sensitive promoter (asr from biobrick BBa_K1231000, an RBS (BBa_B0030) and the Immunity Protein required to neutralise the bacteriocin activity of Colicin Ia (BBa_K1962001).
Usage and Biology
We cloned the Ia-Immunity protein (BBa_K1962001) downstream of a pH sensitive promoter Pasr (BBa_K1231000) and a bile salt sensitive promoter PacrRA (BBa_K1231001) to generate the composite parts (BBa_K1962015) and (BBa_K1962011), respectively.
The immunity proteins for colicin Ia and E3 were cloned in front of the asr pH sensitive promoter. The immunity proteins were amplified with a HA tag fused onto the C-terminal of the immunity proteins to allow for visualisation of expression via western blotting. As shown in Fig 1 we were unable to detect expression of iA-Immunity protein.
Figure 1. Western blot of colicin Ia and E3 immunities with pasr: The col Ia and E3 immunities were amplified with a HA tag to detect expression via a western blot. The asr promoter was blotted alongside as a control. No expression of the immunities in front of the asr promoter was detected.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 215
Illegal AgeI site found at 422 - 1000COMPATIBLE WITH RFC[1000]
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