Difference between revisions of "Part:BBa K1962012"
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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.
 
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.
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Revision as of 20:37, 22 October 2016


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 containing a pSB1C3 backbone. A HA tag was fused onto the C-terminal of the immunity proteins to allow visualisation of expression via western blotting (Fig 1).

IA-e3-acrra.png

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


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 471
    Illegal XhoI site found at 785
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]