Difference between revisions of "Team:Valencia UPV/Improve"

Revision as of 23:15, 9 October 2018

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Part Improvement

The BioBrick chosen to be improved was the YFP coding sequence: BBa_K592101. To do this, we first standardized the part to be compatible with the GoldenBraid 3.0 assembly method creating the part BBa_K2656021. Then, we created the part BBa_K2656020. This part is the CDS of the YFP with the addition of the degradation tag ssRA LVA.

To test the effect of the degradation tag, we designed an experiment with which we measured the increase in protein degradation due to this tag. To perform this experiment, we assembled two composite parts with the same promoter, RBS and terminator:

BBa_K2656112:

BBa_K2656004: the J23106 promoter in its GoldenBraid compatible version from our Part Collection
BBa_K2656009: the B0030 ribosome biding site in its GoldenBraid compatible version from our Part Collection
BBa_K2656021: The original part BBa_K592101 compatible with the GoldenBraid assembly method
BBa_K2656026: the B0015 transcriptional terminator in its GoldenBraid compatible version from our Part Collection

BBa_K2656111:

BBa_K2656004
BBa_K2656009
BBa_K2656020: The original part with the added LVA ssRA degradation tag compatible with the GoldenBraid assembly method
BBa_K2656026

Once the experiment was carried out, the results were plotted and Figure 1 was obtained, in which we can observe that the growth of the bacteria with both constructions was very similar, while the fluorescence had a clear variation.

Figure 1. Experimental results of the fluorescence comparison experiment between the transcriptional unit with BBa_K2656021 and the one with BBa_K2656020.

These data were optimized with our model and the parameters from Table 1 were obtained. With these parameters it is possible to obtain that the protein degradation of the protein with the LVA degradation tag is around twice as much as without the LVA tag.

Table 1. Optimized values of translation rate, degradation rate and dilution rate from experimental data

Optimized parameters

Values

Translation rate p

For YFP with LVA tag BBa_K2656020: p = 0.4622 min^-1
For YFP without LVA tag BBa_K2656021: p = 1.053 min^-1

PoI degradation rate d_p

For YFP with LVA tag BBa_K2656020: d_p = 0.01492 min^-1
For YFP without LVA tag BBa_K2656021: d_p = 0.0080 min^-1

Dilution rate μ

For YFP with LVA tag BBa_K2656020: μ = 0.01166 min^-1
For YFP without LVA tag BBa_K2656021: μ = 0.01307 min^-1

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-<p>These data were optimized with our <a href="https://2018.igem.org/Team:Valencia_UPV/Model#const_models"> model</a> and the parameters from Table 1 were obtained. With these parameters it is possible to obtain that <b>the protein degradation of the protein with the degradation tag is around twice as much as without the tag</b>.
+<p>These data were optimized with our <a href="https://2018.igem.org/Team:Valencia_UPV/Model#const_models"> model</a> and the parameters from Table 1 were obtained. With these parameters it is possible to obtain that <b>the protein degradation of the protein with the LVA degradation tag is around twice as much as without the LVA tag</b>.
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+<a href="https://static.igem.org/mediawiki/2018/7/73/T--Valencia_UPV--dp_ratioUPV2018.png" data-lightbox="true">
+						<img src="https://static.igem.org/mediawiki/2018/7/73/T--Valencia_UPV--dp_ratioUPV2018.png" />
+					</a>
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