Coding

Part:BBa_K2355002

Designed by: Philip SÝrensen   Group: iGEM17_DTU-Denmark   (2017-10-23)


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.

Figure 1: The picture to the left is of amilCP with His-tag spun down in LB media.
The picture to the right is a picture of amilCP with His-tag after the lysis procedure by sonication.


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.

Figure 2: 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.

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Categories
Parameters
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