iGEM 2018 — Team BUCT-China

[![](https://static.igem.org/mediawiki/2018/8/8b/HQ_page_logo.jpg)](https://2018.igem.org)

# Team:BUCT-China/Notebook

![](https://static.igem.org/mediawiki/2018/4/4f/2018_igem_mobile_menu_logo.svg)

![](https://static.igem.org/mediawiki/igem.org/3/3b/Menu_icon.svg)

[![](https://static.igem.org/mediawiki/2018/9/9b/T--BUCT-China--xiaohui.jpeg)](https://2018.igem.org/Team%3ABUCT-China)
[HOME](https://2018.igem.org/Team%3ABUCT-China)

TEAM

[Team Members](https://2018.igem.org/Team%3ABUCT-China/Team)
[Collaborations](https://2018.igem.org/Team%3ABUCT-China/Collaborations)

PROJECT

[Description](https://2018.igem.org/Team%3ABUCT-China/Description)
[Design](https://2018.igem.org/Team%3ABUCT-China/Design)
[Experiments](https://2018.igem.org/Team%3ABUCT-China/Experiments)
[Notebook](https://2018.igem.org/Team%3ABUCT-China/Notebook)
[InterLab](https://2018.igem.org/Team%3ABUCT-China/InterLab)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Results](https://2018.igem.org/Team%3ABUCT-China/Results)
[Demonstrate](https://2018.igem.org/Team%3ABUCT-China/Demonstrate)
[Improve](https://2018.igem.org/Team%3ABUCT-China/Improve)
[Attributions](https://2018.igem.org/Team%3ABUCT-China/Attributions)

PARTS

[Parts Overview](https://2018.igem.org/Team%3ABUCT-China/Parts)
[Basic Parts](https://2018.igem.org/Team%3ABUCT-China/Basic_Part)
[Composite Parts](https://2018.igem.org/Team%3ABUCT-China/Composite_Part)
[Part Collection](https://2018.igem.org/Team%3ABUCT-China/Part_Collection)

[SAFETY](https://2018.igem.org/Team%3ABUCT-China/Safety)

HUMAN PRACTICES

[Human Practices](https://2018.igem.org/Team%3ABUCT-China/Human_Practices)
[Education & Engagement](https://2018.igem.org/Team%3ABUCT-China/Public_Engagement)

AWARDS

[Applied Design](https://2018.igem.org/Team%3ABUCT-China/Applied_Design)
[Entrepreneurship](https://2018.igem.org/Team%3ABUCT-China/Entrepreneurship)
[Hardware](https://2018.igem.org/Team%3ABUCT-China/Hardware)
[Measurement](https://2018.igem.org/Team%3ABUCT-China/Measurement)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Plant](https://2018.igem.org/Team%3ABUCT-China/Plant)
[Software](https://2018.igem.org/Team%3ABUCT-China/Software)

[JUDGING FORM ⇗](https://igem.org/2018_Judging_Form?team=BUCT-China)

# Notebook

2017.12

We designed 4 sets of plasmids and selected [pBbE8a-RFP] plasmid as the master plate for transformation. The No. 1, No. 3, and No. 4 plasmids were loaded with fatty acid operons, glyoxylate operons and another fatty acid operons, respectively. The No. 2 plasmid was used as a vector plasmid to replace the white segment with the arabinose operon, which later reached the role of the vector plasmid. After that, we sent the designed four sets of plasmid information to addgen for plasmid synthesis.

2018.1

We transformed the plasmid into competent cells, cultured and proliferated, and preserved. The plasmid was extracted and subjected to DNA electrophoresis verification, that is, qualitative verification was performed.

![](https://static.igem.org/mediawiki/2018/thumb/9/9c/T--BUCT-China--home7.png/798px-T--BUCT-China--home7.png)

Next, the plasmid was digested. At the start and end of our operon, there were Bgl Ⅱ and Xho Ⅰ cleavage sites, respectively. Therefore, the corresponding restriction enzyme was used for digestion. After that, the enzyme-cut products were subjected to DNA electrophoresis, and each set of plasmids was cut into two sections, which corresponded to two strips on the electrophoresis gel on the electrophoresis gel. Thereafter, a gel recovery operation was carried out, and for the first, third, and fourth groups of plasmids, a band having a molecular weight of about 1,700 was recovered, which was the intended operon.

![](https://static.igem.org/mediawiki/2018/thumb/1/19/T--BUCT-China--home8.png/798px-T--BUCT-China--home8.png)

For the second group, a band having a molecular weight of about 4,000 was recovered to serve as a carrier. The recovered plasmid fragments are then ligated by ligating the target operons obtained in groups 1, 3, and 4 with the vector plasmid fragments of the two groups to form a new plasmid. The next step is to transfer the target plasmid to competent bacteria for a new round of plasmid amplification.

We extracted the plasmids in bacteria No. 1, 3 and 4, and cut the plasmid with the enzymes Bgl II and Xho I, and then verified by electrophoresis.It was found that all the molecular weights in 1, 3 and 4 are divided into two strips ,the molecular weight of a strip is 1700 and another one is 4000. It can be verified that our previous digestion and plasmid ligation operations are correct.

2018.2

We verified the function of the operon, and set up 4 sets of control experiments for each plasmid. The specific experimental conditions are as follows:

* Arabinose: Add 20% arabinose solution
* Fatty acid: Add 100mmol/L fatty acid solution
* Glyoxylic acid: Add 2mol/L glyoxylic acid solution

![](https://static.igem.org/mediawiki/2018/a/ad/T--BUCT-China--result_11.png)

### Detailed information about the January and February experiment can be found in the documentation: [BUCT--China--Notebook.](https://static.igem.org/mediawiki/2018/1/16/T--BUCT-China--Notebook.docx)

2018.7

In July, We performed a fluorescence test on the plasmid and completed the Interlab experiment.

Fluorescence detection of fatty acid operons, we want to use the E. coli knockout of the fade gene to accurately detect the fluorescence value, but unfortunately, our knockout and reconstruction work has not been completed, so our original bacteria measured fluorescence value For details, please refer to:[BUCT-China/Experiments](https://2018.igem.org/Team%3ABUCT-China/Experiments) .

For the fluorescence detection of the glyoxylic acid operon, we used the plasmid that has been successfully constructed and introduced into E. coli for quantitative determination. The data showed that the glyoxylic acid operon played a role. For details, please see:[BUCT-China/Experiments](https://2018.igem.org/Team%3ABUCT-China/Experiments) .

More information about Interlab can be found in [Interlab--BUCT-China](https://2018.igem.org/Team%3ABUCT-China/InterLab) .


[![](https://static.igem.org/mediawiki/2018/8/8b/HQ_page_logo.jpg)](https://2018.igem.org)

# Team:BUCT-China/Experiments

![](https://static.igem.org/mediawiki/2018/4/4f/2018_igem_mobile_menu_logo.svg)

![](https://static.igem.org/mediawiki/igem.org/3/3b/Menu_icon.svg)

[![](https://static.igem.org/mediawiki/2018/9/9b/T--BUCT-China--xiaohui.jpeg)](https://2018.igem.org/Team%3ABUCT-China)
[HOME](https://2018.igem.org/Team%3ABUCT-China)

TEAM

[Team Members](https://2018.igem.org/Team%3ABUCT-China/Team)
[Collaborations](https://2018.igem.org/Team%3ABUCT-China/Collaborations)

PROJECT

[Description](https://2018.igem.org/Team%3ABUCT-China/Description)
[Design](https://2018.igem.org/Team%3ABUCT-China/Design)
[Experiments](https://2018.igem.org/Team%3ABUCT-China/Experiments)
[Notebook](https://2018.igem.org/Team%3ABUCT-China/Notebook)
[InterLab](https://2018.igem.org/Team%3ABUCT-China/InterLab)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Results](https://2018.igem.org/Team%3ABUCT-China/Results)
[Demonstrate](https://2018.igem.org/Team%3ABUCT-China/Demonstrate)
[Improve](https://2018.igem.org/Team%3ABUCT-China/Improve)
[Attributions](https://2018.igem.org/Team%3ABUCT-China/Attributions)

PARTS

[Parts Overview](https://2018.igem.org/Team%3ABUCT-China/Parts)
[Basic Parts](https://2018.igem.org/Team%3ABUCT-China/Basic_Part)
[Composite Parts](https://2018.igem.org/Team%3ABUCT-China/Composite_Part)
[Part Collection](https://2018.igem.org/Team%3ABUCT-China/Part_Collection)

[SAFETY](https://2018.igem.org/Team%3ABUCT-China/Safety)

HUMAN PRACTICES

[Human Practices](https://2018.igem.org/Team%3ABUCT-China/Human_Practices)
[Education & Engagement](https://2018.igem.org/Team%3ABUCT-China/Public_Engagement)

AWARDS

[Applied Design](https://2018.igem.org/Team%3ABUCT-China/Applied_Design)
[Entrepreneurship](https://2018.igem.org/Team%3ABUCT-China/Entrepreneurship)
[Hardware](https://2018.igem.org/Team%3ABUCT-China/Hardware)
[Measurement](https://2018.igem.org/Team%3ABUCT-China/Measurement)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Plant](https://2018.igem.org/Team%3ABUCT-China/Plant)
[Software](https://2018.igem.org/Team%3ABUCT-China/Software)

[JUDGING FORM ⇗](https://igem.org/2018_Judging_Form?team=BUCT-China)

# Experiments

In order to test whether the operons we designed can perform the expected results, we first performed a computer simulation of the protein structure to preliminarily verify the function of the operon. Then,the operon is synthesized into the plasmid backbone by endonucleases and ligases.Next,the plasmid was introduced into TOP 10 competent cells, and the operon was tested for the specific function by detecting the fluorescence of E. coli.Finally,we quantitatively add the inducer to the strain culture solution to quantitatively measure the effect of the inducer concentration on the expression of transcriptional regulatory factors.

### Computer simulation

* Look up data in genebank
* Design our Parts
* Simulate the function and structure of the target protein by using amber software

![](https://static.igem.org/mediawiki/2018/5/50/T--BUCT-China--homeAllR.png)

### Synthesis of plasmid

* Entrusted company to synthesize plasmid backbone and destination sequence
* Cleavage of the sequence and plasmid backbone with restriction endonucleases named BglⅡ and XhoⅠ
* Use DNA ligase to join the sequence to the plasmid backbone to synthesize complete Parts .
  The arabinose operon on the vector plasmid is located upstream of the operon of interest, thereby conferring viability on the operon of interest. At the same time, the AMP resistance gene on the vector plasmid facilitates subsequent screening of the target colony

![](https://static.igem.org/mediawiki/2018/thumb/1/1b/T--BUCT-China--meiqie.jpg/180px-T--BUCT-China--meiqie.jpg)

### Preliminarily verify the function of the operon

* Principle : Both the arabinose operon and our target operon produce a repressor protein, and the repressor protein binds to the operon.[1] When RNA polymerase is transcribed, it is blocked by the repressor protein, and transcription cannot be performed, thereby inhibiting transcription. Taking the fatty acid operon as an example, when we add arabinose, arabinose binds to the repressor protein on the arabinose operon and causes it to fall off. RNA polymerase is slid through the arabinose operon and slid to the downstream fatty acid operon to be repressed by a repressor protein on the fatty acid operon. At this time, we add fatty acids, and the fatty acids bind to the repressor protein, causing it to fall off. RNA polymerase continues to slide and slides over the fluorescent protein gene downstream of the operon to express the fluorescent protein.
* Conversion：Introducing plasmid into competent cells
* Set different inducer gradients
Arabinose: Add 200 μL of 5% arabinose solution
Fatty acid: add 100 μL of 100mmol/L fatty acid solution
Glyoxylic acid: Add 100 μL of 2mol/L glyoxylic acid solution
![](https://static.igem.org/mediawiki/2018/a/ad/T--BUCT-China--result_11.png)* Observe the fluorescence concentration in the test tube every half hour

![](https://static.igem.org/mediawiki/2018/b/bb/T--BUCT-China--result3.jpeg)
![](https://static.igem.org/mediawiki/2018/2/2c/T--BUCT-China--result4.jpeg)
![](https://static.igem.org/mediawiki/2018/4/40/T--BUCT-China--result7.jpeg)

Detailed information about the experiment of "Preliminarily verify the function of the operon"can be found in [Notebook-BUCT-China.](https://2018.igem.org/Team%3ABUCT-China/Notebook)

#### Results:

Fatty acid ec operon:Fluorescent protein expression of No. 1 tube and No. 2 tube, no fluorescence of No. 3 tube and No. 4 tube.
Glyoxylic acid operon:No. 1 tube and No. 4 tube have no fluorescence, and No. 2 tube and No. 3 tube have fluorescent expression
Fatty acid bh operon:No fluorescent expression was observed in No. 1 tube, No. 2 tube, No. 3 tube, and No. 4 tube.

More information about the Results can be found in [Results--BUCT-China](https://2018.igem.org/Team%3ABUCT-China/Results) .

### Quantitatively verify the function of the operon

* Principle : The experimental principle is basically the same as the qualitative experiment.
   The difference is that we can establish the mathematical model by measuring the fluorescence value of the culture under the same conditions of the OD value of each culture.Fluorescence value is proportional to the relationship between time.
* Experimental protocol: For the fatty acid operon, we designed four control groups to measure the fluorescence value. The first group only added arabinose; the second group added different concentrations of fatty acid after adding arabinose for one hour; The group only added gradient fatty acids; the fourth group was a blank control group.
* Stage results: The data on the right is the fluorescence value measured for the first three hours after eight hours of bacterial culture. From this set of data, it can be seen that the fluorescence value of the second group is higher than that of the first group and the blank control group, indicating that the fatty acid operon plays a role.
* Follow-up experiment: We want to knock out the Fade gene in E. coli in subsequent experiments to eliminate the effect of this gene on the detection of fluorescence. This experiment is in progress.

![](https://static.igem.org/mediawiki/2018/d/d7/T--BUCT-China--fashuju.png)

* Principle : The experimental principle is basically the same as the qualitative experiment.
   The difference is that we can establish the mathematical model by measuring the fluorescence value of the culture under the same conditions of the OD value of each culture.Fluorescence value is proportional to the relationship between time.
* Experimental protocol: For the glyoxylic acid operon, we designed four sets of control experiments to measure the fluorescence value, the first group only added arabinose; the second group added glyoxylic acid after one hour of arabinose reaction; the third group B Aldehydic acid; the fourth group was a blank control group.
* Stage results: The data on the right is the fluorescence value measured for the first three hours after eight hours of bacterial culture. From this set of data, it can be seen that the fluorescence value of the second group is higher than that of the first group and the blank control group, indicating that the glyoxylic acid operon plays a role.

![](https://static.igem.org/mediawiki/2018/d/d6/T--BUCT-China--ga.png)

## Citation

[1]Linda J. Wallace,Gary Wilcox. Regulation of the L-arabinose operon in strains of Escherichia coli containing ColE1-ara hybrid plasmids[J]. MGG Molecular & General Genetics,1979,173(3).

[![](https://static.igem.org/mediawiki/2018/8/8b/HQ_page_logo.jpg)](https://2018.igem.org)

# Team:BUCT-China/Results

![](https://static.igem.org/mediawiki/2018/4/4f/2018_igem_mobile_menu_logo.svg)

![](https://static.igem.org/mediawiki/igem.org/3/3b/Menu_icon.svg)

[![](https://static.igem.org/mediawiki/2018/9/9b/T--BUCT-China--xiaohui.jpeg)](https://2018.igem.org/Team%3ABUCT-China)
[HOME](https://2018.igem.org/Team%3ABUCT-China)

TEAM

[Team Members](https://2018.igem.org/Team%3ABUCT-China/Team)
[Collaborations](https://2018.igem.org/Team%3ABUCT-China/Collaborations)

PROJECT

[Description](https://2018.igem.org/Team%3ABUCT-China/Description)
[Design](https://2018.igem.org/Team%3ABUCT-China/Design)
[Experiments](https://2018.igem.org/Team%3ABUCT-China/Experiments)
[Notebook](https://2018.igem.org/Team%3ABUCT-China/Notebook)
[InterLab](https://2018.igem.org/Team%3ABUCT-China/InterLab)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Results](https://2018.igem.org/Team%3ABUCT-China/Results)
[Demonstrate](https://2018.igem.org/Team%3ABUCT-China/Demonstrate)
[Improve](https://2018.igem.org/Team%3ABUCT-China/Improve)
[Attributions](https://2018.igem.org/Team%3ABUCT-China/Attributions)

PARTS

[Parts Overview](https://2018.igem.org/Team%3ABUCT-China/Parts)
[Basic Parts](https://2018.igem.org/Team%3ABUCT-China/Basic_Part)
[Composite Parts](https://2018.igem.org/Team%3ABUCT-China/Composite_Part)
[Part Collection](https://2018.igem.org/Team%3ABUCT-China/Part_Collection)

[SAFETY](https://2018.igem.org/Team%3ABUCT-China/Safety)

HUMAN PRACTICES

[Human Practices](https://2018.igem.org/Team%3ABUCT-China/Human_Practices)
[Education & Engagement](https://2018.igem.org/Team%3ABUCT-China/Public_Engagement)

AWARDS

[Applied Design](https://2018.igem.org/Team%3ABUCT-China/Applied_Design)
[Entrepreneurship](https://2018.igem.org/Team%3ABUCT-China/Entrepreneurship)
[Hardware](https://2018.igem.org/Team%3ABUCT-China/Hardware)
[Measurement](https://2018.igem.org/Team%3ABUCT-China/Measurement)
[Model](https://2018.igem.org/Team%3ABUCT-China/Model)
[Plant](https://2018.igem.org/Team%3ABUCT-China/Plant)
[Software](https://2018.igem.org/Team%3ABUCT-China/Software)

[JUDGING FORM ⇗](https://igem.org/2018_Judging_Form?team=BUCT-China)

# Results

### Result prediction

Before the formal verification experiment, we first predicted the experimental results based on the experimental principle, and predicted as follows:

![](https://static.igem.org/mediawiki/2018/8/80/T--BUCT-China--result2.jpeg)

The upper part of the white line drawn by the red line is the arabinose operon, and the red part below is our target operon. Both the arabinose operon and our target operon produce a repressor protein, and the repressor protein binds to the operon. When RNA polymerase is transcribed, it is blocked by the repressor protein, and transcription cannot be performed, thereby inhibiting transcription. Taking the fatty acid operon as an example, when we add arabinose, arabinose binds to the repressor protein on the arabinose operon and causes it to fall off. RNA polymerase is slid through the arabinose operon and slid to the downstream fatty acid operon to be repressed by a repressor protein on the fatty acid operon. At this time, we add fatty acids, and the fatty acids bind to the repressor protein, causing it to fall off. RNA polymerase continues to slide and slides over the fluorescent protein gene downstream of the operon to express the fluorescent protein. Going back to the different conditions of each group before, we predict that if our operons function normally, then the No. 1 tube and No. 4 tube of each group must not change color, and the No. 2 tube must be discolored. By consulting the literature, we found that Arabia The sugar operon has a chance to fail, so the tube 3 may change color.

### Qualitative experiment results

* Arabinose: Add 200μL of 5% arabinose solution
* Fatty acid: add 100 μL of 100mmol/L fatty acid solution
* Glyoxylic acid: Add 100 μL of 2mol/L glyoxylic acid solution

![](https://static.igem.org/mediawiki/2018/a/ad/T--BUCT-China--result_11.png)

# Experimental results and analysis

In the experiment, a total of three groups of operons were qualitatively tested. The experimental results are as follows:

### Fatty acid ec operon:

Fluorescent protein expression of No. 1 tube and No. 2 tube, no fluorescence of No. 3 tube and No. 4 tube.

![](https://static.igem.org/mediawiki/2018/b/bb/T--BUCT-China--result3.jpeg)

#### analysis:

Combined with the experimental conditions of the fatty acid ec operon validation, we made the following analysis of the experimental results:
Based on previous experimental results, we conclude that the No. 1 tube incorporates arabinose, which opens the arabinose-controlled transcriptional pathway, but, according to our experiments, in the presence of our fatty acid ec operon repressor The No. 1 tube does not contain fatty acids to open the transcription channel controlled by the fatty acid ec operon, so transcription cannot proceed normally, and there should be no fluorescent expression, but the experimental results are fluorescent expression, thus indicating our fatty acid ec operon It did not function as expected, so it was judged that the fatty acid ec operon failed. The failure of the fatty acid ec operon can explain the results of the remaining three tubes: the addition of arabinose to tube 2 opens the transcriptional channel controlled by the arabinose operon, while the fatty acid ec operon fails, transcription proceeds normally, and fluorescence is expressed. No. 3 tube. In tube 4, no arabinose was added, and the transcription channel was controlled by the arabinose operon. Transcription was impossible and no fluorescence was expressed. The results were inconsistent with the expectation, indicating that the fatty acid ec operon failed.

### Glyoxylic acid operon

No. 1 tube and No. 4 tube have no fluorescence, and No. 2 tube and No. 3 tube have fluorescent expression

![](https://static.igem.org/mediawiki/2018/2/2c/T--BUCT-China--result4.jpeg)

#### analysis:

Combined with experimental predictions, the following conclusions were drawn: No. 1 tube added only arabinose, which opened a transcriptional channel that may be controlled by the arabinose operon, but did not add glyoxylate, so it failed to open the transcription controlled by the glyoxylate operon. Channel, so transcription is not working properly, and tube 1 is not fluorescent. The No. 2 tube was added with arabinose and glyoxylate, which respectively opened the transcription channel controlled by the arabinose operon and the glyoxylate operon, so that transcription was normally carried out and fluorescently expressed. Neither the No. 3 tube nor the No. 4 tube was added with arabinose, and the transcription channel controlled by the arabinose operon could not be opened, so that no fluorescent expression was observed. The results are consistent with the predictions, indicating that the glyoxylic acid operon experiment was successful.

### Fatty acid bh operon:

No fluorescent expression was observed in No. 1 tube, No. 2 tube, No. 3 tube, and No. 4 tube.

![](https://static.igem.org/mediawiki/2018/2/2c/T--BUCT-China--result4.jpeg)

#### analysis:

The results of this experiment are very strange. After discussion, our team came to the conclusion that, in view of the addition of arabinose and fatty acids in the No. 2 tube (opening the transcriptional channels controlled by the arabinose operon and the glyoxylate operon, respectively), There was still no fluorescence expression, while the No. 1 tube added only arabinose (opening the transcriptional channel controlled by the arabinose operon). We hypothesized that the fatty acid bh operon has the function of controlling (repressing) the transcriptional pathway, but the substrate (fatty acid) we added is not the correct inducer and therefore cannot open the transcriptional channel controlled by the fatty acid bh operon. We added another substrate to test the fatty acid bh operon and the fatty acid ec operon that had not previously worked.

## Supplementary experiment

Change the substrate: fatty acid for hydroxy fatty acid to test fatty acid ec, fatty acid bh operon

### Experimental results and analysis

### Fatty acid ec operon:

Our main concern is the No. 1 tube, No. 3 tube, No. 5 tube and No. 6 tube (corresponding to the No. 1 tube, No. 2 tube, No. 3 tube, No. 4 tube of the previous experiment). No. 1 tube had fluorescence expression, No. 3 tube had fluorescent expression, and No. 5 tube and No. 6 tube had no fluorescence expression.

![](https://static.igem.org/mediawiki/2018/4/40/T--BUCT-China--result7.jpeg)

#### analysis:

The experimental results are consistent with the expectation that the No. 1 tube only added arabinose, which opened the transcription channel that may be controlled by the arabinose operon, but did not add glyoxylate, so it failed to open the transcription channel controlled by the glyoxylate operon. Transcription was not normal and No. 1 tube was not fluorescent. The No. 3 tube was added with arabinose and glyoxylate, which respectively opened the transcription channel controlled by the arabinose operon and the glyoxylate operon, so that transcription was normally carried out and fluorescently expressed. Neither the No. 5 tube nor the No. 6 tube was added with arabinose, and the transcription channel controlled by the arabinose operon could not be opened, so that no fluorescent expression was observed. The results are consistent with the expectation that the hydroxy fatty acid is a suitable substrate for the fatty acid ec operon and successfully validated our fatty acid ec operon.

### Fatty acid bh operon:

None of the 6 tubes showed fluorescence.

![](https://static.igem.org/mediawiki/2018/f/f2/T--BUCT-China--result8.jpeg)

#### analysis:

Based on an analysis of the previous experimental results for the fatty acid bh operon, we can conclude that hydroxy fatty acids are not a suitable substrate for the induction of the fatty acid bh operon and, therefore, the fatty acid bh operon experiment failed.

### Qualitative analysis total results:

The effectiveness of the fatty acid ec operon and the glyoxylate operon was successfully verified, and the induced substrates were hydroxy fatty acid and glyoxylic acid, respectively. Therefore, based on the success of the experimental results, we have uploaded our fatty acid ec operon and glyoxylate operon (as composite part numbers: BBa\_K2782012 and BBa\_K2782011).

![](https://static.igem.org/mediawiki/2018/8/85/T--BUCT-China--result9.jpeg)

Fatty acid ec operon(to upload as a composite part number: BBa\_K2782012)

![](https://static.igem.org/mediawiki/2018/8/8e/T--BUCT-China--result10.jpeg)

Glyoxylic acid operon (to upload as a composite part number: BBa\_K2782011)

View the corresponding composite parts

* [Fatty acid ec operon](http://parts.igem.org/Part%3ABBa_K2782012)
* [Glyoxylic acid operon](http://parts.igem.org/Part%3ABBa_K2782011)

### Quantitatively verify the results of the operon:

Fluorescence value is proportional to the relationship between time.

Experimental protocol: For the fatty acid operon, we designed four control groups to measure the fluorescence value. The first group only added arabinose; the second group added different concentrations of fatty acid after adding arabinose for one hour; The group only added gradient fatty acids; the fourth group was a blank control group.

Stage results: The data on the right is the fluorescence value measured for the first three hours after eight hours of bacterial culture. From this set of data, it can be seen that the fluorescence value of the second group is higher than that of the first group and the blank control group, indicating that the fatty acid operon plays a role.

Follow-up experiment: We want to knock out the Fade gene in E. coli in subsequent experiments to eliminate the effect of this gene on the detection of fluorescence. This experiment is in progress.

![](https://static.igem.org/mediawiki/2018/d/d7/T--BUCT-China--fashuju.png)

Experimental protocol: For the glyoxylic acid operon, we designed four sets of control experiments to measure the fluorescence value, the first group only added arabinose; the second group added glyoxylic acid after one hour of arabinose reaction; the third group B Aldehydic acid; the fourth group was a blank control group.

Stage results: The data on the right is the fluorescence value measured for the first three hours after eight hours of bacterial culture. From this set of data, it can be seen that the fluorescence value of the second group is higher than that of the first group and the blank control group, indicating that the glyoxylic acid operon plays a role.

![](https://static.igem.org/mediawiki/2018/d/d6/T--BUCT-China--ga.png)