iGEM 2018 — Team KUAS_Korea

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

# Team:KUAS Korea/Notebook

# Notebook

Document the dates you worked on your project. This should be a detailed account of the work done each day for your project.

### What should this page have?

* Chronological notes of what your team is doing.
* Brief descriptions of daily important events.
* Pictures of your progress.
* Mention who participated in what task.

### Inspiration

You can see what others teams have done to organize their notes:

* [2014 ATOMS-Turkiye](https://2014.igem.org/Team%3AATOMS-Turkiye/Notebook)
* [2014 Tec Monterrey](https://2014.igem.org/Team%3ATec-Monterrey/ITESM14_project.html#tab_notebook)
* [2014 Kyoto](https://2014.igem.org/Team%3AKyoto/Notebook/Magnetosome_Formation#title)
* [2014 Cornell](https://2014.igem.org/Team%3ACornell/notebook)


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

# Team:KUAS Korea/Experiments

# [KUAS](/Team%3AKUAS_Korea)

* Team
  + [Team](/Team%3AKUAS_Korea/Team)
  + [Attributions](/Team%3AKUAS_Korea/Attributions)
  + [Collaborations](/Team%3AKUAS_Korea/Collaborations)
  + [Official Team Profile](//igem.org/Team.cgi?team_id=2564)
* Project
  + [Description](/Team%3AKUAS_Korea/Description)
  + [Design](/Team%3AKUAS_Korea/Design)
  + [Parts](/Team%3AKUAS_Korea/Parts)
  + [Achievement](/Team%3AKUAS_Korea/Project/Achievement)
* Wet Lab
  + [Experiments](/Team%3AKUAS_Korea/Experiments)
  + [Results](/Team%3AKUAS_Korea/Results)
  + [InterLab](/Team%3AKUAS_Korea/InterLab)
  + [Safety](/Team%3AKUAS_Korea/Safety)
* Dry Lab
  + [Modeling](/Team%3AKUAS_Korea/Model)
* Human Practices
  + [Overview](/Team%3AKUAS_Korea/HumanPractices/Overview)
  + [Human Practices](/Team%3AKUAS_Korea/HumanPractices/HumanPractices)
  + [Integrated Practices](/Team%3AKUAS_Korea/HumanPractices/IntegratedPractices)
  + [Public Engagement](/Team%3AKUAS_Korea/Public_Engagement)
* [Notebook](/Team%3AKuas_Korea/Notebook)

|  |
| --- |
| Experiments |

#### **1. Designing and constructing the cooperator and cheater**

We have looked the previous iGEM projects and articles on the game theory. We decided to express GFP for cheater and beta-glucosidase for cooperator. We received the designed constitutive expression vector applicable to ligation independent cloning using the plasmid containing BBa\_J23106 from Ph.D. Ko.

#### **2.Transformation into E.coli BW25113**

As the *E.coli* BW25113 is a host strain for the surface display and expression, we transformed the designed plasmid vector into it. We cultured the bacteria on M9 minimal media; one with glucose and another with cellobiose respectively. We checked the activity of cheater on M9 minimal medium with glucose and cooperator on M9 minimal medium with cellobiose.

#### **3.Co-culture of cooperator and cheater with different ratio**

1. Both cooperator and cheater were incubated in M9 minimal medium with 0.48% glucose at 37’C until it reached 1.0 in OD 600.
2. Batch co-culture was conducted in 5ml of M9 minimal medium with 0.48% cellobiose solution; in ratio of cheater and cooperator 1:1, 1:2, 1:4, 2:1, 4:1.
3. After 2, 4 and 6 days, each co-culture medium with different ratio was extracted and streaked on LB agar plate with ampicillin.
4. After overnight growth, total cell was counted and the plate was checked under fluorescence microscopy to count cheater; GFP expressing bacteria.

### Sponsors

![](https://static.igem.org/mediawiki/2013/1/13/Spon2.png)
![](https://static.igem.org/mediawiki/2016/f/f4/Korea_U_Seoul_Sponsor_ku.gif)
![](https://static.igem.org/mediawiki/2013/1/1c/Spon3.png)
![](https://static.igem.org/mediawiki/2016/3/34/Korea_U_Seoul_sponsor_kusso.jpg)
![](https://static.igem.org/mediawiki/2018/8/8b/HQ_page_logo.jpg)

### KUAS

Korea University

iGEM

#### Links

* [Home](/Team%3AKUAS_Korea)
* [Project](/Team%3AKUAS_Korea/Description)
* [Team](/Team%3AKUAS_Korea/Team)
* [Human Practices](/Team%3AKUAS_Korea/Human_Practices)
* [FACEBOOK](//facebook.com/KUAS2018iGEM)

#### About Us

**Student Leader:** Taehoon Lee
**Wet Lab Leader:** Bosun Ryu
**Dry Lab Leader:** Uijin Han
**Human Practices Leader:** Eunmin Kim

#### Contact Us

Seoul Seongbuk-gu Anam-ro 145
**Tel:** 02)1234-5678
**Email:** kuaskorea@gmail.com

© Copyright **KUAS 2018**. All Rights Reserved

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

# Team:KUAS Korea/Results

# [KUAS](/Team%3AKUAS_Korea)

* Team
  + [Team](/Team%3AKUAS_Korea/Team)
  + [Attributions](/Team%3AKUAS_Korea/Attributions)
  + [Collaborations](/Team%3AKUAS_Korea/Collaborations)
  + [Official Team Profile](//igem.org/Team.cgi?team_id=2564)
* Project
  + [Description](/Team%3AKUAS_Korea/Description)
  + [Design](/Team%3AKUAS_Korea/Design)
  + [Parts](/Team%3AKUAS_Korea/Parts)
  + [Achievement](/Team%3AKUAS_Korea/Project/Achievement)
* Wet Lab
  + [Experiments](/Team%3AKUAS_Korea/Experiments)
  + [Results](/Team%3AKUAS_Korea/Results)
  + [InterLab](/Team%3AKUAS_Korea/InterLab)
  + [Safety](/Team%3AKUAS_Korea/Safety)
* Dry Lab
  + [Modeling](/Team%3AKUAS_Korea/Model)
* Human Practices
  + [Overview](/Team%3AKUAS_Korea/HumanPractices/Overview)
  + [Human Practices](/Team%3AKUAS_Korea/HumanPractices/HumanPractices)
  + [Integrated Practices](/Team%3AKUAS_Korea/HumanPractices/IntegratedPractices)
  + [Public Engagement](/Team%3AKUAS_Korea/Public_Engagement)
* [Notebook](/Team%3AKuas_Korea/Notebook)

|  |
| --- |
| Results |

#### **1. Beta-glucosidase expression**

Cooperator expresses beta-glucosidase with constitutive promoter to degrade cellobiose around it. We had the cell stock of cooperator with pCEL vector containing beta-glucosidase and transformed it into E.coli BW25113. The medium plate contains cellobiose instead of glucose to check whether E.coli that we transformed expresses beta-glucosidase on its surface. The streaking side below shows that cooperator grows on cellobiose containing M9 minimal medium.

![](https://static.igem.org/mediawiki/2018/0/0c/T--KUAS_Korea--BETAGlucosidase_expression.png)

[Fig 1.] Cultured cooperator on M9 minimal medium with 0.48% cellobiose

#### **2. GFP expression**

Cheater expresses GFP when it obtains glucose. We had the cell stock of cheater with the vector encoding GFP as a reporter gene and transformed it firstly into E.coli DH5α and then to E.coli BW25113.

![](https://static.igem.org/mediawiki/2018/4/48/T--KUAS_Korea--GFP_PART1.png)

[Fig 2.] Cultured cheater on M9 minimal medium with 0.48% glucose

![](https://static.igem.org/mediawiki/2018/1/1b/T--KUAS_Korea--GFP_PART2.png)

[Fig 2-1.] Cultured cheater under fluorescence microscopy showing the GFP activity

#### **3. Co-culture**

As the cheater and cooperator reached OD600 1.8 in M9 minimal medium with glucose concentration 0.48%, we began co-culture in M9 minimal medium with cellobiose concentration 0.48%. Batch co-culture was conducted in 5ml of medium in ratio of cheater and cooperator 1:1, 1:2, 1:4, 2:1, 4:1. 1 was counted as 1ul and each medium was incubated for 1, 3 and 4 days. After each day, we streaked 100ul of the incubated medium on LB agar plate with ampicillin. However, expected bacteria did not grow at all after overnight incubation.

We predicted possible causes of unsuccessful co-culture as follows;

* Too little amount of bacteria inoculated in the medium
* Low survival rate of pre-cultured bacteria since the optimal OD600 was 1.0 and ours was 1.8 which is thought to be after stationary phase.
* Too short period of incubation

We plan to conduct the experiment once again with newly grown bacteria.

### Sponsors

![](https://static.igem.org/mediawiki/2013/1/13/Spon2.png)
![](https://static.igem.org/mediawiki/2016/f/f4/Korea_U_Seoul_Sponsor_ku.gif)
![](https://static.igem.org/mediawiki/2013/1/1c/Spon3.png)
![](https://static.igem.org/mediawiki/2016/3/34/Korea_U_Seoul_sponsor_kusso.jpg)
![](https://static.igem.org/mediawiki/2018/8/8b/HQ_page_logo.jpg)

### KUAS

Korea University

iGEM

#### Links

* [Home](/Team%3AKUAS_Korea)
* [Project](/Team%3AKUAS_Korea/Description)
* [Team](/Team%3AKUAS_Korea/Team)
* [Human Practices](/Team%3AKUAS_Korea/Human_Practices)
* [FACEBOOK](//facebook.com/KUAS2018iGEM)

#### About Us

**Student Leader:** Taehoon Lee
**Wet Lab Leader:** Bosun Ryu
**Dry Lab Leader:** Uijin Han
**Human Practices Leader:** Eunmin Kim

#### Contact Us

Seoul Seongbuk-gu Anam-ro 145
**Tel:** 02)1234-5678
**Email:** kuaskorea@gmail.com

© Copyright **KUAS 2018**. All Rights Reserved