Difference between revisions of "Part:BBa C0060:Experience"

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===Applications of BBa_C0060===
 
===Applications of BBa_C0060===
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The characterization of BBa_C0060 was carried out, first, by creating a composite part (BBa_K2471000) capable of expressing the biopart of interest; this was done by the addition of a T7 promoter and RBS (BBa_K525998) and a T1 terminator (BBa_B0010) to the basic part. The design was first done in silico utilizing SnapGene®, where the program indicated that the plasmid would have a length of 3,010 base pairs. After verifying the feasibility of creating this BioBrick®, the part was synthesized using the 3A assembly method. Agarose gel (1%) electrophoresis was performed to corroborate that the obtained molecular weight coincided with the expected one.
  
 
===User Reviews===
 
===User Reviews===

Revision as of 20:42, 1 November 2017

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_C0060

The characterization of BBa_C0060 was carried out, first, by creating a composite part (BBa_K2471000) capable of expressing the biopart of interest; this was done by the addition of a T7 promoter and RBS (BBa_K525998) and a T1 terminator (BBa_B0010) to the basic part. The design was first done in silico utilizing SnapGene®, where the program indicated that the plasmid would have a length of 3,010 base pairs. After verifying the feasibility of creating this BioBrick®, the part was synthesized using the 3A assembly method. Agarose gel (1%) electrophoresis was performed to corroborate that the obtained molecular weight coincided with the expected one.

User Reviews

UNIQ4fae13bacd611920-partinfo-00000000-QINU

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UNIPV-Pavia iGEM 2011

NB: unless differently specified, all tests were performed in E. coli MGZ1 in M9 supplemented medium at 37°C in low copy plasmid pSB4C5.

The AiiA enzyme activity has been characterized under the regulation of ptet promoter, assaying its enzymatic activity, with these parts:


A system of differential equations has been derived and parameters would have been identified, studying the exponential growth phase:
The four measurment parts ptet-RBSx-LuxI were quantitatively characetrized using the BBa_T9002 biosensor. Unfortunately the model parameters (kcat, kM,AiiA) were not estimated because placing the device in low copy plasmid (pSB4C5)resulted in no HSL degradation, as shown in the figures below:
A semi-quantitative information about AiiA enzyme activity is given, performing an experiment in E. coli TOP10, with the four measurement devices in pSB1A2; the results are summerized in the graphs below:
In this way a measure of AiiA activity is given, as the percentage of HSL degraded in 7 hours (measurements provided as average [stdandard deviation]):
Measurement device HSL degraded percentage
BBa_K516220 96.11 [3.66]
BBa_K516221 77.60 [2.66]
BBa_K516222 50.66 [16.87]
BBa_K516224 100 [0]

In order to evaluate the RBSs efficiency, the ratio between the percentage of the degraded HSL realtive to the measurement device with the standard RBS (BBa_B0034) was computed:
RBS RBS efficiency
BBa_B0030 0.96
BBa_B0031 0.78
BBa_B0032 0.51
BBa_B0034 1

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