Team:HokkaidoU Japan/Results

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Results

Until the Wiki freeze, we tried to construct plasmid with the insert we designed and managed to combine plasmid with INSERT 1, 2 or 3(we couldn't insert INSERT 4, which is a control of the INSERT 2 or 3).

To assay our construct, we cultured E. coli with IPTG induction, purified the crude extract by the His-tag and condensed them using

When the expressed protein (both INSERT 2 and 3) were cut at the signal peptides, the molecular weight becomes 52367.76 or 48575.69 respectively.

The figure bellow is the result of SDS-PAGE. We applied purified INSERT 2 (left) and INSERT 3 (right, not visible)and found that the band of INSERT 2 is at the correct place.

We assumed two possibility that explain why we couldn't get the band of INSERT 3; the concentration of protein is too small, the His-tag didn't interacted with the Nickel column and flowed through.

To determine the reason why we couldn't see the band, we conducted phytase activity assay with the crude extract, purified and condensed products, and the flow through of purification process.

As the table and figure above shows, we can see high phytase activity in the flow through and concluded that His-tag didn't interacted with the Nickel, probably because the His-tag was not exposed outside. Simultaneously, we can assume that self-Assembling Peptides helped His-tag being exposed and interacting with Nickel.

We applied crude extract of INSERT 2 (left) and INSERT 3 (right),

Here you can describe the results of your project and your future plans.

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• Clearly and objectively describe the results of your work.
• Future plans for the project.
• Considerations for replicating the experiments.

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• Interpretation of the results obtained during your project. Don't just show a plot/figure/graph/other, tell us what you think the data means. This is an important part of your project that the judges will look for.
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