iGEM 2020 — Team CAU_China

[![](https://static.igem.org/mediawiki/2020/8/8b/HQ_page_logo.jpg)](https://2020.igem.org)

# Team:CAU China/Notebook

* [Home](https://2020.igem.org/Team%3ACAU_China)
* [Model](https://2020.igem.org/Team%3ACAU_China/Model)
* [Team](https://2020.igem.org/Team%3ACAU_China/Team%20Member)
  + [Team Member](https://2020.igem.org/Team%3ACAU_China/Team%20Member)
  + [Attribution](https://2020.igem.org/Team%3ACAU_China/Attribution)
  + [Contribution](https://2020.igem.org/Team%3ACAU_China/Contribution)
* [Experiment](https://2020.igem.org/Team%3ACAU_China/Design)
  + [Design](https://2020.igem.org/Team%3ACAU_China/Design)
  + [Implementation](https://2020.igem.org/Team%3ACAU_China/Implementation)
  + [Engineering](https://2020.igem.org/Team%3ACAU_China/Engineering)
  + [Proof of Concept](https://2020.igem.org/Team%3ACAU_China/Proof%20of%20Concept)
  + [Notebook](https://2020.igem.org/Team%3ACAU_China/Notebook)
  + [Protocol](https://2020.igem.org/Team%3ACAU_China/Protocol)
* [Project](https://2020.igem.org/Team%3ACAU_China/Description)
  + [Description](https://2020.igem.org/Team%3ACAU_China/Description)
  + [Part](https://2020.igem.org/Team%3ACAU_China/Part)
  + [Safety Form](https://2020.igem.org/Team%3ACAU_China/Safety%20Form)
* [Education & Science Communication](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Overview)
  + [Overview](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Overview)
  + [Synthetic Biology from Scratch](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/SyntheticBiologyfromScratchonlinelivelesson0)
  + [Take you to synthetic biology](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Takeyoutosyntheticbiology2)
  + [Meetings](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Meetings3)
  + [Others](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Others4)
* [Human Practice](https://2020.igem.org/Team%3ACAU_China/Overview)
  + [Overview](https://2020.igem.org/Team%3ACAU_China/Overview)
  + [Integrated Human Practice](https://2020.igem.org/Team%3ACAU_China/Integrated%20Human%20Pratice)
* [Collaboration & Partnership](https://2020.igem.org/Team%3ACAU_China/Collaboration)
  + [Collaboration](https://2020.igem.org/Team%3ACAU_China/Collaboration)
  + [Partnership](https://2020.igem.org/Team%3ACAU_China/Partnership)

![](https://static.igem.org/mediawiki/2020/f/f6/T--CAU_China--notebooktop.jpg)

[August](#General)
[September](#Plasmids)
[October](#t3)

# August

August 24thAugust 25thAugust 26thAugust 27thAugust 28thAugust 29thAugust 30th

# September

September 1stSeptember 2ndSeptember 3rd September 12th September 13thSeptember 14thSeptember 18thSeptember 19thSeptember 21th September 22th September 23th September 24th September 25thSeptember 26thSeptember 27thSeptember 30th

# October

October 1stOctober 2ndOctober 3rdOctober 4thOctober 5th October 6th
 October 9th October 10th October 11thOctober 12th October 13th October 14thOctober 16thOctober 17th October 22th October 26th

[![](https://static.igem.org/mediawiki/2020/8/8b/HQ_page_logo.jpg)](https://2020.igem.org)

# Team:CAU China/Engineering

* [Home](https://2020.igem.org/Team%3ACAU_China)
* [Model](https://2020.igem.org/Team%3ACAU_China/Model)
* [Team](https://2020.igem.org/Team%3ACAU_China/Team%20Member)
  + [Team Member](https://2020.igem.org/Team%3ACAU_China/Team%20Member)
  + [Attribution](https://2020.igem.org/Team%3ACAU_China/Attribution)
  + [Contribution](https://2020.igem.org/Team%3ACAU_China/Contribution)
* [Experiment](https://2020.igem.org/Team%3ACAU_China/Design)
  + [Design](https://2020.igem.org/Team%3ACAU_China/Design)
  + [Implementation](https://2020.igem.org/Team%3ACAU_China/Implementation)
  + [Engineering](https://2020.igem.org/Team%3ACAU_China/Engineering)
  + [Proof of Concept](https://2020.igem.org/Team%3ACAU_China/Proof%20of%20Concept)
  + [Notebook](https://2020.igem.org/Team%3ACAU_China/Notebook)
  + [Protocol](https://2020.igem.org/Team%3ACAU_China/Protocol)
* [Project](https://2020.igem.org/Team%3ACAU_China/Description)
  + [Description](https://2020.igem.org/Team%3ACAU_China/Description)
  + [Part](https://2020.igem.org/Team%3ACAU_China/Part)
  + [Safety Form](https://2020.igem.org/Team%3ACAU_China/Safety%20Form)
* [Education & Science Communication](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Overview)
  + [Overview](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Overview)
  + [Synthetic Biology from Scratch](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/SyntheticBiologyfromScratchonlinelivelesson0)
  + [Take you to synthetic biology](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Takeyoutosyntheticbiology2)
  + [Meetings](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Meetings3)
  + [Others](https://2020.igem.org/Team%3ACAU_China/ScienceCommunication%26Education_/Others4)
* [Human Practice](https://2020.igem.org/Team%3ACAU_China/Overview)
  + [Overview](https://2020.igem.org/Team%3ACAU_China/Overview)
  + [Integrated Human Practice](https://2020.igem.org/Team%3ACAU_China/Integrated%20Human%20Pratice)
* [Collaboration & Partnership](https://2020.igem.org/Team%3ACAU_China/Collaboration)
  + [Collaboration](https://2020.igem.org/Team%3ACAU_China/Collaboration)
  + [Partnership](https://2020.igem.org/Team%3ACAU_China/Partnership)

![](https://static.igem.org/mediawiki/2020/2/24/T--CAU_China--engineeringtop.jpg)

## Engineering Success

## Our Design

To design a dsRNA part which we can control its expression, we need an induction factor. However, for most of the inducible promoter, it requires huge amount of inducer when we apply for factory manufacture for farmers. In addition, many of the inducers are chemical toxicity to engineering bacteria/fungi. So we searched for an inducer which is low-toxicity to our yeast. We then found light-inducible protein is a good choice.

For light inducible protein, it has following advantages: 1. Fast and efficient response to light signals; 2. Having no extra affect to strains.

We searched and sorted our result about light-control system and finally chose CarH protein as our optogenetic automatic regulation system which has a induction fold of 350 (To learn more, please click to our design page).

In order to achieve our goal of manipulating yeast synthesizing dsRNA, we inserted CarO operon in our dsRNA sequence upstream. As CarH tetramer binding with the CarO DNA operon, RNA polymerase would be hindered by CarH protein for steric hindrance. dsRNA wouldn't be produced. When the light on, CarH tetramer would break into monomers as Co-C bound photolysis. Then the yeast can produce dsRNA.

With optogenetic automated regulation system, we can achieve the goal of controlling dsRNA synthesis. Here we'll elaborate our part building, testing and learning.

## Optogenetic Automated Regulation system

We design an optogenetic automated regulation system to help us produce dsRNA. Automated optogenetic control system is divided into two modules:1. yeast growth system;2. dsRNA synthesis system. The strain grows to ideal population to OD600 0.5~0.6. Then the strain transfer to dsRNA synthesis system. In this system, dsRNA is transcribed and measured by pepper RNA. When dsRNA meets the ideal concentration, yeast strain will be treated as ready to use dried inactivated yeast powder.

To achieve engineering success, we will introduce two part into our engineering *Saccromyces cerevisiae*: CarH Light control system and dsRNA manufacture system. We will use *auco troph* yeast ΔTrp, ΔLeu with PGBKT7 and PGADT7 plasmid construct our system.
We will build our system with restriction enzyme cutting and combining. Then we will introduce our constructed plasmid into our engineering yeast with electrotransformation and chemical transformation.
After we transformed the construct into yeast, we will use bacterial PCR to identify transformed yeast.

Then we'll incubate transformed yeast in different temperature in order to get CarH protein. We will use SDS-PAGE assay and western-blot to test our result.

After we transform the yeast successfully which can express CarH protein, we'll transform our dsRNA part. We'll then screen transformed engineering yeast with SC-Dropout medium. After we identify transformed yeast, we'll incubate yeast in dark and turn 525nm light on when the yeast population meets to target value. We'll produce dsRNA in 525nm light environment and measure the concentration of dsRNA by using HBC fluorescent.

![](https://static.igem.org/mediawiki/2020/c/c5/T--CAU_China--Flow_Chart.jpg)

[![](https://static.igem.org/mediawiki/2020/8/8b/HQ_page_logo.jpg)](https://2020.igem.org)

# Team:CAU China/Experiments

# Experiments

Describe the research, experiments, and protocols you used in your iGEM project. These should be detailed enough for another team to repeat your experiments.

If you made Parts this year, please remember to put all information, characterization, and measurement data on the Part's Main Page on the [Registry](http://parts.igem.org/Main_Page).

### What should this page contain?

* Protocols
* Experiments

### Inspiration

* [2014 Colombia](https://2014.igem.org/Team%3AColombia/Protocols)
* [2014 Imperial](https://2014.igem.org/Team%3AImperial/Protocols)
* [2014 Caltech](https://2014.igem.org/Team%3ACaltech/Project/Experiments)

[![](https://static.igem.org/mediawiki/2020/8/8b/HQ_page_logo.jpg)](https://2020.igem.org)

# Team:CAU China/Results

# Results

You can describe the results of your project and your future plans here.

### What should this page contain?

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

### Describe what your results mean

* 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.
* Show data, but remember **all measurement and characterization data must also be on the Part's Main Page on the [Registry](http://parts.igem.org/Main_Page).** Otherwise these data will not be in consideration for any medals or part awards!
* Consider including an analysis summary section to discuss what your results mean. Judges like to read what you think your data means, beyond all the data you have acquired during your project.

### Project Achievements

You can also include a list of bullet points (and links) of the successes and failures you have had over your summer. It is a quick reference page for the judges to see what you achieved during your summer.

* A list of linked bullet points of the successful results during your project
* A list of linked bullet points of the unsuccessful results during your project. This is about being scientifically honest. If you worked on an area for a long time with no success, tell us so we know where you put your effort.

### Inspiration

See how other teams presented their results.

* [2014 TU Darmstadt](https://2014.igem.org/Team%3ATU_Darmstadt/Results/Pathway)
* [2014 Imperial](https://2014.igem.org/Team%3AImperial/Results)
* [2014 Paris Bettencourt](https://2014.igem.org/Team%3AParis_Bettencourt/Results)