Difference between revisions of "Part:BBa K2355002"
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It is clear that the expression of the blue/purple color is very strong. In conclusion the amilCP has been successfully modified by adding a His-tag without compromising the protein.
 
It is clear that the expression of the blue/purple color is very strong. In conclusion the amilCP has been successfully modified by adding a His-tag without compromising the protein.
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To test whether the amilCP with His-tag worked as expected, we purified two solutions of amilCP; one with His-tagged amilCP, and one with just amilCP.  
 
To test whether the amilCP with His-tag worked as expected, we purified two solutions of amilCP; one with His-tagged amilCP, and one with just amilCP.  

Revision as of 23:27, 1 November 2017


AmilCP with 6x His-tag

This is a modification of the AmilCP part from the Uppsala 2011 team (BBa_K592009). It has a 6x His-tag attached to the C-terminal end of the protein

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]



Results of experiments

We aimed to improve the amilCP part from the distribution kit by adding a 6x his-tag to the C-terminal end of the chromoprotein. By doing this, it would theoretically be possible to purify both the amilCP part, and composite parts based on the his-tagged amilCP, using affinity chromatography.

The first picture is amilCP with His-tag spun down in LB media.

T--DTU-Denmark--results-amilcp-assembly-16a.png

The second is a picture of amilCP with His-tag after the lysis procedure by sonication.

T--DTU-Denmark--results-amilcp-assembly-16b.png

It is clear that the expression of the blue/purple color is very strong. In conclusion the amilCP has been successfully modified by adding a His-tag without compromising the protein.

To test whether the amilCP with His-tag worked as expected, we purified two solutions of amilCP; one with His-tagged amilCP, and one with just amilCP.

T--DTU-Denmark--results-amilcp-purification-comparison.png

Figure 3: Comparison of how amilCP with or without His-tag is purified. The data used graph is from the samples with the highest amount of sample loading (5 times). The absorbance was measured at 588 nm.

From these results we can conclude that our his-tag purification of amilCP has worked.