iGEM 2021 — Team Thessaly

[![](https://static.igem.org/mediawiki/2021/8/8b/HQ_page_logo.jpg)](https://2021.igem.org)

# Team:Thessaly/Notebook

[![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--amalth-logo.png)](https://2021.igem.org/Team%3AThessaly)

* [Home](https://2021.igem.org/Team%3AThessaly)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Communication)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

×

* [Home](#hero)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Education)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

FlexStart Bootstrap Template - Index

Notebook

* [Start](#1)
* [February](#2)
* [March](#3)
* [April](#4)
* [May](#5)
* [June](#6)
* [July](#7)
* [August](#8)
* [September](#9)
* [October](#10)
* [November](#11)

## Overview

This year marks the 3rd consecutive year the team participated in the competition and
the second year of our project Amalthea. Even though Covid-19 prohibited us from meeting
in person in the first half we were finally granted permission to meet in person and continue
our work hand in hand (not literally though).

## Timeline

Start

#####

-iGEM Thessaly 2021 - First Meeting, the first time the teammates are
introduced to each other with the help of our wonderful advisors.

![](https://static.igem.org/mediawiki/2021/3/39/T--Thessaly--f-meeting.jpg)

February

#####

-Research on past iGEM projects and synthetic biology lectures.
-Getting to know the team members over the internet and planning
our meetings and schedule

March

#####

- Brainstorming sessions about ways we can improve on last years project
- First Dry lab meeting with advisors
- Our Fundraising journey just began
- The wet lab team met for the first time in person
- The whole team had a brainstorming session in Alkazar Park in Larissa.
- We presented our project to the Hellenic Medical Students' International Committee.

![](https://static.igem.org/mediawiki/2021/5/55/T--Thessaly--brnstrm.png)

-We were part of the Athens Science Festival and through a video and
crossword puzzle introduced our project.

![](https://static.igem.org/mediawiki/2021/1/1a/T--Thessaly--athscnfestival.png)

-We met with the PI’s to inform them about our project progress and get some feedback

April

#####

-We attended the International Forum for Medical Students and
Junior Doctors to inform medical students about our project and how
impactful it can be.

![](https://static.igem.org/mediawiki/2021/9/9c/T--Thessaly--medicalstd.jpg)

-We had our first integrated meeting with a company that produces probiotics and we discussed a potential partnership.

![](https://static.igem.org/mediawiki/2021/4/40/T--Thessaly--notebook1.png)

-We met with another scientific team from NTUA(National Technical University of Athens) to get an insight of how the team works to find sponsorships.

May

#####

-We had our first meeting with the team that we partnered with, which is the Aalto-Helsinki team and it was lovely.
-We had an integrated meeting with Dr. Spahos that was really influential to the development of our application.
-Finished the modeling of the lac operon module, the most complex part of our in-silico design.
-Another important meeting concerning the modelling of our project with Dr. Stamatakis.
-Our first meeting with the McMaster Synbio to talk about future collaborations as we have many similarities in our projects.
-We held an event in the central square of Larissa on the topic of gut health. This was our first in person event we organized.

![](https://static.igem.org/mediawiki/2021/e/e0/T--Thessaly--notebook3.jpg)

June

#####

-We had another really important meeting about our modeling with
-Dr. Klapa that left us wanting more time with her…
-We filmed our promotional video and we successfully uploaded it to iGEM video Universe!
-Panel discussion hosted by us and McMaster Synbio with the title “New frontiers in Gut Research”

July

#####

-Integrated Human Practices Meeting with Dr. Klapa
-We were guests on the Aalto-Helsinki podcast.
-We had an in person integrated meeting with a fish farm and we went back and forth on the use of probiotics used in fish farming.
-We were guests to the annual Greek Meet-Up
-Dry Lab Safety and Security Workshop hosted by us and Aalto-Helsinki team with the goal of informing our team members about our concerns and goals.

![](https://static.igem.org/mediawiki/2021/b/b2/T--Thessaly--notebook4.png)

-We attended the podcast of iGEM Patra

August

#####

-We made a presentation to the scouts of Greece about SDG's
-We successfully completed our whole modeling (whoooo!)
-We met with the Aalto-Helsinki to plan a potential jamboree party!!
-We started writing this page! (meta)
-We hosted with the collaboration of the Aalto-Helsinki team the MicrobioCOSMOS event on the 20th of this month and it was awesome you can see the presentations from the event here
(link [here](https://www.youtube.com/watch?v=FmgFjxboVMU&t=1s))

![](https://static.igem.org/mediawiki/2021/d/d1/T--Thessaly--notebook2.jpg)

-We held an outdoor event by the name “Microbiome: Learn and Create”
for children to promote our project and synthetic biology through
mutual learning and outdoor activities.

![](https://static.igem.org/mediawiki/2021/e/e9/T--Thessaly--notebook5.png)

September

#####

-We had a meeting in which we discussed with a large pharmaceutical company about the perspectives of our project as a product
-We hosted a seminar and presented our school guides to high school teachers
-We conducted an Integrated Human Practices interview at a farm and at a veterinarian office

![](https://static.igem.org/mediawiki/2021/c/cf/T--Thessaly--timeline-img4.jpg)

October

#####

-We filmed our presentation video
-We hosted an event about the Mediterranean diet named “My Mediterranean diet”
-We organized a reading of our book for children to a kids’ theatrical group

November

#####

-We had our online judging session on November 6th and met up with other iGem teams at Brussels.

![](https://static.igem.org/mediawiki/2021/5/51/T--Thessaly--fly-thessaly-logo.png)

[![](https://static.igem.org/mediawiki/2021/9/9c/T--Thessaly--facebookColoredResized..png)](https://www.facebook.com/igemthessaly/)

[![](https://static.igem.org/mediawiki/2021/a/a2/T--Thessaly--instagramLogo-Resized.png)](https://www.instagram.com/igemthessaly/?hl=el)

[![](https://static.igem.org/mediawiki/2021/e/ef/T--Thessaly--twitterLogo-Resized.png)](https://twitter.com/igemthessaly?lang=el)

[![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--youtubeLogo-Resized.png)](https://www.youtube.com/channel/UCBHXzFL7r9xxHxqQICznphA)

igem.thessaly@gmail.com

![](https://static.igem.org/mediawiki/2021/7/71/T--Thessaly--PfizerLogo.png)

![](https://static.igem.org/mediawiki/2021/6/6c/T--Thessaly--logo_EPSILON-HEALTH.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--UTHLogo.png)

![](https://static.igem.org/mediawiki/2021/8/86/T--Thessaly--Hnet.com-image.png)

![](https://static.igem.org/mediawiki/2021/a/ae/T--Thessaly--ethniki-asfalistiki.png)

![](https://static.igem.org/mediawiki/2021/7/79/T--Thessaly--ZagorinLogo.png)

![](https://static.igem.org/mediawiki/2021/8/8a/T--Thessaly--AnadrasiLogo.png)

![](https://static.igem.org/mediawiki/2021/7/7d/T--Thessaly--FlemingLogo.png)

![](https://static.igem.org/mediawiki/2021/3/3c/T--Thessaly--Lab_SolutionsLogo.png)

![](https://static.igem.org/mediawiki/2021/5/58/T--Thessaly--MediplanLogo.png)

![](https://static.igem.org/mediawiki/2021/9/94/T--Thessaly--MetrolabLogo.png)

![](https://static.igem.org/mediawiki/2021/f/f6/T--Thessaly--PindosLogo.png)

![](https://static.igem.org/mediawiki/2021/b/ba/T--Thessaly--SnapgeneLogo.png)

![](https://static.igem.org/mediawiki/2021/b/bf/T--Thessaly--SuperchipLogo.png)

![](https://static.igem.org/mediawiki/2021/0/0c/T--Thessaly--idt.png)

![](https://static.igem.org/mediawiki/2021/3/32/T--Thessaly--la-travel.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--twist.png)

[![](https://static.igem.org/mediawiki/2021/8/8b/HQ_page_logo.jpg)](https://2021.igem.org)

# Team:Thessaly/Engineering

[![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--amalth-logo.png)](https://2021.igem.org/Team%3AThessaly)

* [Home](https://2021.igem.org/Team%3AThessaly)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Communication)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

×

* [Home](#hero)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Education)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

FlexStart Bootstrap Template - Index

![](https://static.igem.org/mediawiki/2021/1/1b/T--Thessaly--curve-arrow.png)

Engineering

[Click On The Last Branch](#1)

## Overview

Engineering represents a strategy of problem solving or development of new technologies and products.
Under this perspective, assumptions and approximations have to be expressed through a thorough design
in order to define the system's variables, possible outputs and behaviour under different conditions.
In this page, we show how we performed our system’s modeling, concluded in our final hardware for the
capsule and designed our wet lab experiments concerning both the capsule and the probiotic based on the
DBTL cycle.

GPCR MODULEPFLIC MODULETYROSINE-TYROSINASE MODULESCFAs PRODUCTIONCUMATE KILL-SWITCHEXTRACELLULAR IPTG STABILITYTYROSINASE AS LAC OPERON'S OUTPUTHARDWARE

![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--modal-img4.png)

## Tango GPCR metabolite detection module

![](https://static.igem.org/mediawiki/2021/5/58/T--Thessaly--G1A.png)
**Figure 1:** *The NOT GATE based TANGO GPCR SCFAs detection module, inhibiting the Signal in presence of SCFAs.*

The TANGO GPCR SCFAs detection module is a whole-cell biosensor, hosted inside the tank inside the
[bioelectronic capsule](https://2021.igem.org/Team%3AThessaly/Hardware)
beneath the Semipermeable membrane, that allows SCFAs to pass through. SCFAs activate the GPCR
*FFAR2*  that is fused to a *LacI*  through a linker, containing a TEV Cleavage Site (TCS).
After activation, Beta arrestin *B-arr2* fused to a TEV protease cleaves and releases
*LacI*, suppressing the reporter module. The aforementioned genes encode for eukaryotic
transmembrane proteins that are toxic to bacteria, due to aggregate formation. To overcome this obstacle,
we implemented a genetic circuit that favors bacterial expression of eukaryotic proteins
 (Michou, Stergios & Skretas, 2020).  The final construct([BBa\_K3866014](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866014)) consists of four Transcription
Units (TUs). The three of them are regulated by *araBAD*
[BBa\_K3505025](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505025) ,
[BBa\_K3505026](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505026) ,
[BBa\_K3505032](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505032) , [BBa\_K3505038](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505038) . We had placed the four TUs having reverse orientation in order to
avoid Polymerase slippage using two alpha vectors: α2 [BBa\_K3505009](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505009) and α1 Reverse [BBa\_K3505008](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505008). We
changed the TU's orientation to forward  (Dusek et al., 2020),  solving the problem of DNA loop formation because of the repeated *araBAD* sequences.

### References

1. Dusek, J., Plchova, H., Cerovska, N., Poborilova, Z., Navratil, O., & Kratochvilova, K. et al. (2020). Extended Set of GoldenBraid Compatible Vectors for Fast Assembly of Multigenic Constructs and Their Use to Create Geminiviral Expression Vectors. *Frontiers In Plant Science*, 11. doi: 10.3389/fpls.2020.522059

2. Michou, M., Stergios, A., & Skretas, G. (2020). SuptoxD2.0: A second‐generation engineered Escherichia coli strain achieving further enhanced levels of recombinant membrane protein production. *Biotechnology And Bioengineering*, 117(8), 2434-2445. doi: 10.1002/bit.27378

## Promoter Flic Metabolite detection module

![](https://static.igem.org/mediawiki/2021/c/c7/T--Thessaly--eng-img6.png)
**Figure 1:** *The NOT GATE based pFliC SCFAs detection module, inhibiting the Signal in presence of SCFAs.*

The detection system of the capsule is based on the NOT-GATE device. The signal is in low concentrations of SCFAs. The promoter Flic is sensitive for SCFAs especially for butyrate  (Tobe, Nakanishi and Sugimoto, 2010).  We conducted experiments in order to verify the activation of pFlic
by butyrate and the functionality of our module Level omega:*LacO:ECFP:term-pflic:LacI:term*
([BBa\_K3866036](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866036)). During our experiments, we changed the fluorescent protein *ECFP*
([BBa\_K3866028](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866028)) to *sfGFP* ([BBa\_K3866030](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866030)), a stronger fluorescent protein, in order to be sure
that the signal is not false positive and that the range between negative and positive control is
distinct.

### References

1. Tobe, T., Nakanishi, N. and Sugimoto, N., 2010. Activation of Motility by Sensing Short-Chain Fatty Acids via Two Steps in a Flagellar Gene Regulatory Cascade in Enterohemorrhagic Escherichia coli. *Infection and Immunity*, 79(3), pp.1016-1024.

## Tyrosine / Tyrosinase Reporter module

![](https://static.igem.org/mediawiki/2021/e/ec/T--Thessaly--eng-img2.png)
**Figure 1:** *The arabinose inducible reporter module, Converting L-Tyrosine to L-Dopa and L-Dopa quinone for electrochemical output.*

![](https://static.igem.org/mediawiki/2021/6/6a/T--Thessaly--T1A.png)
**Figure 2:** *The metabolic pathway of L-Tyrosine conversion catalyzed by Tyrosinase for optical output.*

Tyrosinase *Tyr1* is an enzyme responsible for the conversion of L-Tyrosine to L-DOPA.
Tyrosinase is transported to the extracellular membrane by the fused *AIDA* autotransporter
and its signal peptide ([BBa\_K3866010](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866010)), and the catalysis that performs, leads to the subsequent
release of electrons while L-DOPA converts to L-DOPA quinone (VanArsdale, 2020). This electrical current can be
detected and serve as the step of conversion from the biological to the electrochemical signal in
our bioelectronic capsule. The first iteration we performed was choosing an arabinose inducible
promoter *araBAD* ([BBa\_K3866000](http://parts.igem.org/wiki/index.php/Part%3ABBa_K3866000)) for the production of our protein. This helped us overcome
the metabolic strain of constitutive expression ([BBa\_K3505012](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3505012)). In order to make sure that our
system is both induced and sensitive, we also worked with a range of different concentrations
(0,002%, 0,02%, 0,2%, 1%, 2%, 6%, 12%) of the inducer molecule, L-arabinose.

### References

1. VanArsdale, E., Hornstrom, D., Sjoberg, G., Jarbur, I., Pitzer, J., Payne, G. F., … Bentley, W. E. (2020). A co-culture based tyrosine-tyrosinase electrochemical gene circuit for connecting cellular communication with electronic networks. ACS Synthetic Biology. doi:10.1021/acssynbio.9b00469

## SCFAs Production modules

Our proposed solution is a probiotic able to produce the quantities of SCFAs **missing** inside the gut.
We designed the constructs for **Acetate**, **Propionate** & **Butyrate** producing enzymes.
The Acetate production enzymes ***pta*** and ***ack*** catalyze the conversion from Acetyl CoA to
Acetate. Concurrently with Acetate production ATP is produced.
ATP activates the negative feedback loop of which *acs* is responsible
 (Enjalbert, B.et al. 2017).  For high yield production we chose a **Δacs** strain.

![](https://static.igem.org/mediawiki/2021/3/37/T--Thessaly--eng-img9.png)
**Figure 1:** *The arabinose inducible Acetate production module, utilizing AckA and Pta.*

![](https://static.igem.org/mediawiki/2021/8/8c/T--Thessaly--E5.png)
**Figure 2:** *The metabolic pathway of Acetate production.*

The propionate-producing genetic construct ([BBa\_K3866031](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866031)) consists of the three enzymes: Sbm
([BBa\_K3866023](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866023)), YgfG ([BBa\_K3866024](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866024)) & YgfH ([BBa\_K3866025](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866025)). These enzymes catalyze the conversion of
succinyl-CoA into propionate  (Akawi et al, 2015).  For **high yield** production of propionate and
butyrate we chose an **ΔldhA** strain. The lack of ldhA, increases the carbon flux by promoting the
anaerobic metabolism instead of the aerobic one.

![](https://static.igem.org/mediawiki/2021/a/a4/T--Thessaly--E7.png)
**Figure 3:** *The arabinose inducible Propionate production module, utilizing sbm, ygfG and ygfH.*

![](https://static.igem.org/mediawiki/2021/f/f0/T--Thessaly--E8.png)
**Figure 4:** *The metabolic pathway of Propionate production.*

Butyrate is produced by nine enzymes that catalyze the formation of butyrate from acetyl coA.
 (Miscevic, D. et al. 2019). The genes coding for those enzymes were cloned into two operons:
*BktB, Hbd, PhaJ, Ptb, Buk and PhaA, PhaB, Crt and Ter* ([BBa\_K3866015](http://parts.igem.org/wiki/index.php?title=Part:BBa_K3866015)).

![](https://static.igem.org/mediawiki/2021/1/16/T--Thessaly--op1.png)
![](https://static.igem.org/mediawiki/2021/b/b4/T--Thessaly--op2.png)
**Figure 5:** *The arabinose inducible Butyrate production module, utilizing the two operons.*

![](https://static.igem.org/mediawiki/2021/a/a4/T--Thessaly--eng-img11.png)
**Figure 6:** *The metabolic pathway of Butyrate production.*

There were **two main iterations** on the SCFA production modules. The first one was to change the
orientation of our constructs from reverse to forward as we also did for the
Tango module. The second one was to replace our initial promoters with arabinose inducible ones,
*araBAD*, converting our expression system from a constitutive to an inducible one.

### References

1. Akawi L, Srirangan K, Liu X, Moo-Young M, Perry Chou C. (2015) Engineering Escherichia coli for high-level production of propionate. *J Ind Microbiol Biotechno*l. Jul;42(7):1057-72.<https://doi.org/10.1007/s10295-015-1627-4>

2. Enjalbert, B., Millard, P., Dinclaux, M., Portais, J., & Létisse, F. (2017). Acetate fluxes in Escherichia coli are determined by the thermodynamic control of the Pta-AckA pathway. *Scientific Reports*, 7(1). doi: 10.1038/srep42135.

3. Miscevic, D., Srirangan, K., Kefale, T., Abedi, D., Moo-Young, M., & Chou, C. P. (2019). Production of cellulosic butyrate and 3-hydroxybutyrate in engineered Escherichia coli. *Applied microbiology and biotechnology*, 103(13), 5215–5230.<https://doi.org/10.1007/s00253-019-09815-x>

4. Pontrelli, S., Chiu, T. Y., Lan, E. I., Chen, F. Y., Chang, P., & Liao, J. C. (2018). Escherichia coli as a host for metabolic engineering. *Metabolic engineering*, 50, 16–46.<https://doi.org/10.1016/j.ymben.2018.04.008>

## Cumate kill-switch module

![](https://static.igem.org/mediawiki/2021/8/8c/T--Thessaly--eng-img5.png)
**Figure 1:** *The mechanism of cumate gene switch (Seo SO, Schmidt-Dannert C.2019).*

Safety is of major importance for us, considering that we deal with genetically modified bacteria.
That’s why one of our kill-switches focuses on killing the bacteria in a controlled manner even
before exiting the gut. The **cumate inducible kill-switch** consists of a repressor, **CymR**, a cumate operator, **CuO**, and a toxin-antitoxin system, **mazE-mazF**  (Choi Y. et al. 2010).  In the **absence** of
Cumate, CymR binds to the cumate operator and suppresses the toxin mazF. In the **presence** of
Cumate, CymR loses its affinity to CuO, the toxin is expressed and the bacteria die. At first, we
designed our constructs with level 0 interchangeable parts with the GoldenBraid assembly. However,
if we assembled a transcription unit with the level 0 toxin part, our bacteria would die. So, we
redesigned our module in order to assemble all transcription units simultaneously, containing both
the antitoxin and the repressor.

### References

1. Choi, Y. J., Morel, L., Le François, T., Bourque, D., Bourget, L., Groleau, D., Massie, B., & Míguez, C. B. (2010). Novel, versatile, and tightly regulated expression system for Escherichia coli strains. Applied and environmental microbiology, 76(15), 5058–5066. <https://doi.org/10.1128/AEM.00413-10>

2. Seo, S. O., & Schmidt-Dannert, C. (2019). Development of a synthetic cumate-inducible gene expression system for Bacillus. Applied microbiology and biotechnology, 103(1), 303–313.<https://doi.org/10.1007/s00253-018-9485-4>

## Εxtracellular IPTG stability

We started off our modeling from building and simulating the lac operon module, the most complex
and challenging part of our design. It is an IPTG (Isopropyl β-D-1-thiogalactopyranoside) induced
genetic mechanism in which we make a clear distinction between intra- and extracellular IPTG.
Extracellular IPTG is the component that we used to control in order to induce and test the system.
After some experiments that exhibited instability, we realized that extracellular IPTG had to be a
fixed value during the simulations, as it practically is in vitro; by integrating that in our model
and re-running the simulations we ended up having a smoother, more precise outcome.

![](https://static.igem.org/mediawiki/2021/9/9e/T--Thessaly--model-figure-6.png)
**Figure 1:** *The lac operon modified, “open” genetic circuit.*

### References

1. Stamatakis, M., & Mantzaris, N. V. (2009). Comparison of deterministic and stochastic models of the lac operon genetic network. Biophysical Journal, 96(3), 887–906. https://doi.org/10.1016/j.bpj.2008.10.028 <https://doi.org/10.1016/j.bpj.2008.10.028>

## Tyrosine as lac operon’s output

At the first “open circuit” version of lac operon, its output species was GFP, which served as a
general-purpose reporter gene. After looking at some literature, we noticed that GFP’s kinetics
were very hard to depict, since the extraction of a kinetic formula for GFP expression always
results from experimental data fitting and is often characterized by noisy dynamics. This is why we
replaced GFP with Tyrosine. By doing so, we express an output that is chemically compatible with the
Reporter module by using a species that is much more likely to follow the operon’s kinetics. Note
that having GFP as an output did not get in the way of having an effective simulation, but was forcing
the assumption that GFP follows the lac operon’s kinetics, which was rather unrealistic.

![](https://static.igem.org/mediawiki/2021/d/d4/T--Thessaly--model-figure-9.png)
**Figure 1:** *The Reporter Module genetic circuit.*

### References

1. Vanarsdale, E., Hörnström, D., Sjöberg, G., Järbur, I., Pitzer, J., Payne, G. F., Van Maris, A. J. A., & Bentley, W. E. (2020). A coculture based tyrosine-tyrosinase electrochemical gene circuit for connecting cellular communication with electronic networks. ACS Synthetic Biology, 9(5), 1117–1128.<https://doi.org/10.1021/acssynbio.9b00469>

## Hardware

**Overview**
The Design Build Test Learn cycle is embedded in scientific research and naturally was in ours
too. We began this year with an already designed capsule. The goal of this year was to improve
upon that design and implement new ideas that we had. We improved the design by reducing the size
of the capsule and thus making it safer and easier to swallow. Next we implemented our new idea of
data transmittance, which used an SoC. Finally we made one last safety addition to the capsule, by
implementing a mechanical kill switch.

**Iteration 1. Processor and Antenna**
The hardware side of the project is mainly divided into 3 major DBTL (design build test learn) cycles.
The first iteration had to do with the change in the “brains” of our system. Last year we implemented
the processing of our data with a simple PIC18F microprocessor. This year we decided to change that as
we had greater needs in terms of data processing. After diligent research we ended up on the
DA14531 BLE SoC.
This SoC gave us the ability to combine into a single tiny device both the processing and the
transfer of the data as it is equipped with a bluetooth antenna that uses AES/CCM
(Advanced Encryption Standard) encryption to safely transmit the data. In comparison with last year
when we had temporarily implemented a simple RF transmitter, this year we wanted to connect our
capsule to a phone so we had to find a different solution. We concluded on the DA14531 BLE SoC,
which also allowed us to shrink down the capsule.

**Iteration 2. Capsule dimensions**
Taking advantage of the newly available space, our first sub iteration was to add another battery
to the already existing ones, with the goal, of course, to increase the battery life of the system.
The ADC consumes approximately 50 to 60 μA. Our first battery array provided 45mA so our battery
life was approximately 750 hours under ideal conditions. With the third battery we boosted that to
1500 hours. After those iterations we still had space available, so we decided to shrink the capsule
dimensions and specifically on the X axis. The capsule ended up being 15mm in length from the original
22m and with the addition of 1 battery.

![](https://static.igem.org/mediawiki/2021/c/c9/T--Thessaly--end-img7.png)
**Figure 1:** *The 3D design of the capsule showcasing the outer cell and the components*

**Iteration 3. Kill switch**
Finally, we designed a mechanical kill switch that would facilitate the cell death induced by the
biological arabinose inducible kill-switch. This device is located above the bacteria vat and it
gets activated when a certain amount of pressure is applied on the capsule. The amount of pressure required for the activation, is equal to the breaking point that the outer cell of the capsule.
The L-arabinose, that is contained in this separate space, gets mixed with the bacteria, enabling
the expression of the toxin ,and the consequent death of the cells.

### References

1. Zhao, K., Yan, G., Lu, L., & Xu, F. (2015). Low-Power Wireless Electronic Capsule for Long-Term Gastrointestinal Monitoring. Journal of Medical Systems, 39(2). <https://doi.org/10.1007/s10916-015-0211-9>

2. Zhuang, Y., Hou, W., Zheng, X., Wang, Z., Zheng, J., Pi, X., Cui, J., Jiang, Y., Qian, S., & Peng, C. (2011). A MEMS-based electronic capsule for time-controlled drug delivery in the alimentary canal. Sensors and Actuators, A: Physical, 169(1), 211–216. <https://doi.org/10.1016/j.sna.2011.05.024>

![](https://static.igem.org/mediawiki/2021/5/51/T--Thessaly--fly-thessaly-logo.png)

[![](https://static.igem.org/mediawiki/2021/9/9c/T--Thessaly--facebookColoredResized..png)](https://www.facebook.com/igemthessaly/)

[![](https://static.igem.org/mediawiki/2021/a/a2/T--Thessaly--instagramLogo-Resized.png)](https://www.instagram.com/igemthessaly/?hl=el)

[![](https://static.igem.org/mediawiki/2021/e/ef/T--Thessaly--twitterLogo-Resized.png)](https://twitter.com/igemthessaly?lang=el)

[![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--youtubeLogo-Resized.png)](https://www.youtube.com/channel/UCBHXzFL7r9xxHxqQICznphA)

igem.thessaly@gmail.com

![](https://static.igem.org/mediawiki/2021/7/71/T--Thessaly--PfizerLogo.png)

![](https://static.igem.org/mediawiki/2021/6/6c/T--Thessaly--logo_EPSILON-HEALTH.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--UTHLogo.png)

![](https://static.igem.org/mediawiki/2021/8/86/T--Thessaly--Hnet.com-image.png)

![](https://static.igem.org/mediawiki/2021/a/ae/T--Thessaly--ethniki-asfalistiki.png)

![](https://static.igem.org/mediawiki/2021/7/79/T--Thessaly--ZagorinLogo.png)

![](https://static.igem.org/mediawiki/2021/8/8a/T--Thessaly--AnadrasiLogo.png)

![](https://static.igem.org/mediawiki/2021/7/7d/T--Thessaly--FlemingLogo.png)

![](https://static.igem.org/mediawiki/2021/3/3c/T--Thessaly--Lab_SolutionsLogo.png)

![](https://static.igem.org/mediawiki/2021/5/58/T--Thessaly--MediplanLogo.png)

![](https://static.igem.org/mediawiki/2021/9/94/T--Thessaly--MetrolabLogo.png)

![](https://static.igem.org/mediawiki/2021/f/f6/T--Thessaly--PindosLogo.png)

![](https://static.igem.org/mediawiki/2021/b/ba/T--Thessaly--SnapgeneLogo.png)

![](https://static.igem.org/mediawiki/2021/b/bf/T--Thessaly--SuperchipLogo.png)

![](https://static.igem.org/mediawiki/2021/0/0c/T--Thessaly--idt.png)

![](https://static.igem.org/mediawiki/2021/3/32/T--Thessaly--la-travel.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--twist.png)

[![](https://static.igem.org/mediawiki/2021/8/8b/HQ_page_logo.jpg)](https://2021.igem.org)

# Team:Thessaly/Experiments

[![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--amalth-logo.png)](https://2021.igem.org/Team%3AThessaly)

* [Home](https://2021.igem.org/Team%3AThessaly)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Communication)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

×

* [Home](#hero)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Education)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

FlexStart Bootstrap Template - Index

Experiments

* [Protocols](#1)
* [Lab Book](#2)

## Overview

Protocols are the first step of the success of an experiment. This makes them vital for completing the
experimental procedures. So, during the lockdown, we wrote down all the protocols that we should use,
while during our experiments we added all the new protocols we used. Finally, it is worth mentioning
that these protocols were the foundations in order to begin our experiments.

## Protocols

![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--inoue.png)
![](https://static.igem.org/mediawiki/2021/f/fa/T--Thessaly--after-inoue.png)

![](https://static.igem.org/mediawiki/2021/7/7a/T--Thessaly--restriction.png)
![](https://static.igem.org/mediawiki/2021/4/4b/T--Thessaly--after-restriction.png)

![](https://static.igem.org/mediawiki/2021/f/fc/T--Thessaly--one-pot.png)
![](https://static.igem.org/mediawiki/2021/7/75/T--Thessaly--after-one.png)

![](https://static.igem.org/mediawiki/2021/7/74/T--Thessaly--transform.png)
![](https://static.igem.org/mediawiki/2021/d/d5/T--Thessaly--after-transform.png)

![](https://static.igem.org/mediawiki/2021/3/35/T--Thessaly--miniprep.png)
![](https://static.igem.org/mediawiki/2021/e/e7/T--Thessaly--after-miniprep.png)

![](https://static.igem.org/mediawiki/2021/0/01/T--Thessaly--glycerol.png)
![](https://static.igem.org/mediawiki/2021/4/4c/T--Thessaly--after-glycerol.png)

![](https://static.igem.org/mediawiki/2021/b/b5/T--Thessaly--primer.png)
![](https://static.igem.org/mediawiki/2021/2/2b/T--Thessaly--after-prime.png)

![](https://static.igem.org/mediawiki/2021/d/dd/T--Thessaly--pcr-kappa-beforeHover.png)
![](https://static.igem.org/mediawiki/2021/d/d1/T--Thessaly--pcr-kappa-afterHover.png)

![](https://static.igem.org/mediawiki/2021/f/f5/T--Thessaly--q5.png)
![](https://static.igem.org/mediawiki/2021/a/ad/T--Thessaly--after-q5.png)

![](https://static.igem.org/mediawiki/2021/e/ee/T--Thessaly--colony-pcr.png)
![](https://static.igem.org/mediawiki/2021/8/85/T--Thessaly--after-colony.png)

![](https://static.igem.org/mediawiki/2021/d/d9/T--Thessaly--agarose-gel.png)
![](https://static.igem.org/mediawiki/2021/c/ca/T--Thessaly--after-agarose.png)

![](https://static.igem.org/mediawiki/2021/8/85/T--Thessaly--dna-extract.png)
![](https://static.igem.org/mediawiki/2021/2/2a/T--Thessaly--after-dna.png)

![](https://static.igem.org/mediawiki/2021/b/b9/T--Thessaly--before-sds.png)
![](https://static.igem.org/mediawiki/2021/3/36/T--Thessaly--after-sds.png)

![](https://static.igem.org/mediawiki/2021/4/44/T--Thessaly--agar-plates-plain.png)
![](https://static.igem.org/mediawiki/2021/c/c2/T--Thessaly--after-agar.png)

![](https://static.igem.org/mediawiki/2021/4/47/T--Thessaly--lb-liquid.png)
![](https://static.igem.org/mediawiki/2021/2/26/T--Thessaly--after-lb-liquid.png)

![](https://static.igem.org/mediawiki/2021/a/a2/T--Thessaly--nanodrop.png)
![](https://static.igem.org/mediawiki/2021/5/52/T--Thessaly--after-nanodrop.png)

![](https://static.igem.org/mediawiki/2021/e/ee/T--Thessaly--before-alginate.png)
![](https://static.igem.org/mediawiki/2021/b/b6/T--Thessaly--after-alginate.png)

![](https://static.igem.org/mediawiki/2021/a/a5/T--Thessaly--plate.png)
![](https://static.igem.org/mediawiki/2021/f/f7/T--Thessaly--after-plate.png)

![](https://static.igem.org/mediawiki/2021/6/64/T--Thessaly--before-cell.png)
![](https://static.igem.org/mediawiki/2021/4/4f/T--Thessaly--after-cell.png)

![](https://static.igem.org/mediawiki/2021/d/d7/T--Thessaly--commonly.png)
![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--after-commonly.png)

![](https://static.igem.org/mediawiki/2021/3/38/T--Thessaly--antibiotik.png)
![](https://static.igem.org/mediawiki/2021/0/04/T--Thessaly--after-antibiotic.png)

![](https://static.igem.org/mediawiki/2021/1/16/T--Thessaly--tae.png)
![](https://static.igem.org/mediawiki/2021/0/05/T--Thessaly--after-tae.png)

![](https://static.igem.org/mediawiki/2021/a/a6/T--Thessaly--m9-medium.png)
![](https://static.igem.org/mediawiki/2021/7/78/T--Thessaly--after-m9.png)

![](https://static.igem.org/mediawiki/2021/d/d8/T--Thessaly--pbs-buffer.png)
![](https://static.igem.org/mediawiki/2021/7/7d/T--Thessaly--after-pbs.png)

![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--tyrosinas.png)
![](https://static.igem.org/mediawiki/2021/d/d9/T--Thessaly--after-tyrosinas.png)

![](https://static.igem.org/mediawiki/2021/b/bd/T--Thessaly--bglx.png)
![](https://static.igem.org/mediawiki/2021/d/df/T--Thessaly--after-bglx.png)

## Lab Book

Reporting the daily conducted experiments in detail is of great importance as it helps us organize
our experiments, always find our mistakes and eventually keep track of the correct
optimizations/changes in our protocols. It is very important to have all the protocols registered
correctly and in an organized manner, in order to be able to work more efficiently and record all
the modifications made to the protocols throughout the wet lab experience. You can explore our Lab
Book to see how it works.

![](https://static.igem.org/mediawiki/2021/5/54/T--Thessaly--lab-book-plain-before.png)
![](https://static.igem.org/mediawiki/2021/e/ed/T--Thessaly--lab-book-hover.png)

![](https://static.igem.org/mediawiki/2021/5/51/T--Thessaly--fly-thessaly-logo.png)

[![](https://static.igem.org/mediawiki/2021/9/9c/T--Thessaly--facebookColoredResized..png)](https://www.facebook.com/igemthessaly/)

[![](https://static.igem.org/mediawiki/2021/a/a2/T--Thessaly--instagramLogo-Resized.png)](https://www.instagram.com/igemthessaly/?hl=el)

[![](https://static.igem.org/mediawiki/2021/e/ef/T--Thessaly--twitterLogo-Resized.png)](https://twitter.com/igemthessaly?lang=el)

[![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--youtubeLogo-Resized.png)](https://www.youtube.com/channel/UCBHXzFL7r9xxHxqQICznphA)

igem.thessaly@gmail.com

![](https://static.igem.org/mediawiki/2021/7/71/T--Thessaly--PfizerLogo.png)

![](https://static.igem.org/mediawiki/2021/6/6c/T--Thessaly--logo_EPSILON-HEALTH.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--UTHLogo.png)

![](https://static.igem.org/mediawiki/2021/8/86/T--Thessaly--Hnet.com-image.png)

![](https://static.igem.org/mediawiki/2021/a/ae/T--Thessaly--ethniki-asfalistiki.png)

![](https://static.igem.org/mediawiki/2021/7/79/T--Thessaly--ZagorinLogo.png)

![](https://static.igem.org/mediawiki/2021/8/8a/T--Thessaly--AnadrasiLogo.png)

![](https://static.igem.org/mediawiki/2021/7/7d/T--Thessaly--FlemingLogo.png)

![](https://static.igem.org/mediawiki/2021/3/3c/T--Thessaly--Lab_SolutionsLogo.png)

![](https://static.igem.org/mediawiki/2021/5/58/T--Thessaly--MediplanLogo.png)

![](https://static.igem.org/mediawiki/2021/9/94/T--Thessaly--MetrolabLogo.png)

![](https://static.igem.org/mediawiki/2021/f/f6/T--Thessaly--PindosLogo.png)

![](https://static.igem.org/mediawiki/2021/b/ba/T--Thessaly--SnapgeneLogo.png)

![](https://static.igem.org/mediawiki/2021/b/bf/T--Thessaly--SuperchipLogo.png)

![](https://static.igem.org/mediawiki/2021/0/0c/T--Thessaly--idt.png)

![](https://static.igem.org/mediawiki/2021/3/32/T--Thessaly--la-travel.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--twist.png)

[![](https://static.igem.org/mediawiki/2021/8/8b/HQ_page_logo.jpg)](https://2021.igem.org)

# Team:Thessaly/Results

[![](https://static.igem.org/mediawiki/2021/c/cb/T--Thessaly--amalth-logo.png)](https://2021.igem.org/Team%3AThessaly)

* [Home](https://2021.igem.org/Team%3AThessaly)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Communication)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

×

* [Home](#hero)
* Project
  + [Description](https://2021.igem.org/Team%3AThessaly/Description)
  + [Engineering](https://2021.igem.org/Team%3AThessaly/Engineering)
  + [Safety](https://2021.igem.org/Team%3AThessaly/Safety)
  + [Proof of Concept](https://2021.igem.org/Team%3AThessaly/Proof_Of_Concept)
  + [Contribution](https://2021.igem.org/Team%3AThessaly/Contribution)
  + [Results](https://2021.igem.org/Team%3AThessaly/Results)
* Human Practices
  + [Integrated](https://2021.igem.org/Team%3AThessaly/Human_Practices)
  + [Education & Engagement](https://2021.igem.org/Team%3AThessaly/Education)
  + [Collaborations](https://2021.igem.org/Team%3AThessaly/Collaborations)
  + [Mental Health](https://2021.igem.org/Team%3AThessaly/Excellence)
  + [Sustainability](https://2021.igem.org/Team%3AThessaly/Sustainable)
  + [Animal Welfare](https://2021.igem.org/Team%3AThessaly/Animal_Welfare)
* Wet Lab
  + [Experiments](https://2021.igem.org/Team%3AThessaly/Experiments)
  + [Parts](https://2021.igem.org/Team%3AThessaly/Parts)
  + [Improvement](https://2021.igem.org/Team%3AThessaly/Improve)
* Dry Lab
  + [Model](https://2021.igem.org/Team%3AThessaly/Model)
  + [Hardware](https://2021.igem.org/Team%3AThessaly/Hardware)
  + [Application](https://2021.igem.org/Team%3AThessaly/Software)
* Product Development
  + [Entrepreneurship](https://2021.igem.org/Team%3AThessaly/Entrepreneurship)
  + [Implementation](https://2021.igem.org/Team%3AThessaly/Implementation)
* Team
  + [Members](https://2021.igem.org/Team%3AThessaly/Team)
  + [Attributions](https://2021.igem.org/Team%3AThessaly/Attributions)
  + [Partnership](https://2021.igem.org/Team%3AThessaly/Partnership)
  + [Notebook](https://2021.igem.org/Team%3AThessaly/Notebook)
  + [Medal Criteria](https://2021.igem.org/Team%3AThessaly/Medal_Criteria)
* [JAMBOREE](https://2021.igem.org/Team%3AThessaly/Giant_Jamboree)

FlexStart Bootstrap Template - Index

Results

[Strategy](#2)

[Capsule](#3)

[Probiotic](#4)

## Overview

The bioelectronic capsule’s and the engineered probiotic’s modules were tested both from a dry and a wet
lab perspective. Based on the module that is tested each time, we divided all the experiments into two
big categories: The detection system that comprises of the pFlic, the TANGO and the Tyrosine/Tyrosinase
module and the amelioration system that comprises of the SCFA production modules, the cellulose
degradation module, the cumate inducible kill-switch and the microencapsulation technique for the
probiotic.

## Strategy

Our team built new genetic modules combining basic DNA parts in a standardized way. In order to increase
the multigene assembly efficiency, we followed the GoldenBraid assembly strategy that is used for Plant
Synthetic Biology and can also facilitate heterologous expression of proteins in bacteria. This enables
us to test our system’s modules separately based on the modularity of our design.

## CAPSULE

Metabolite Detection

GPCR

Read More

Metabolite Detection

pFLiC

Read More

Signal Production

Tyrosinase

Read More

Dry Lab

Model

Read More

## PROBIOTIC

SCFAs production

Acetate

Read More

SCFAs production

Propionate

Read More

SCFAs production

Butyrate

Read More

Cellulose Utilization

BglX

Read More

Administration and Protection

Alginate Beads

Read More

Kill Switch

CymR

Read More

![](https://static.igem.org/mediawiki/2021/5/51/T--Thessaly--fly-thessaly-logo.png)

[![](https://static.igem.org/mediawiki/2021/9/9c/T--Thessaly--facebookColoredResized..png)](https://www.facebook.com/igemthessaly/)

[![](https://static.igem.org/mediawiki/2021/a/a2/T--Thessaly--instagramLogo-Resized.png)](https://www.instagram.com/igemthessaly/?hl=el)

[![](https://static.igem.org/mediawiki/2021/e/ef/T--Thessaly--twitterLogo-Resized.png)](https://twitter.com/igemthessaly?lang=el)

[![](https://static.igem.org/mediawiki/2021/c/c8/T--Thessaly--youtubeLogo-Resized.png)](https://www.youtube.com/channel/UCBHXzFL7r9xxHxqQICznphA)

igem.thessaly@gmail.com

![](https://static.igem.org/mediawiki/2021/7/71/T--Thessaly--PfizerLogo.png)

![](https://static.igem.org/mediawiki/2021/6/6c/T--Thessaly--logo_EPSILON-HEALTH.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--UTHLogo.png)

![](https://static.igem.org/mediawiki/2021/8/86/T--Thessaly--Hnet.com-image.png)

![](https://static.igem.org/mediawiki/2021/a/ae/T--Thessaly--ethniki-asfalistiki.png)

![](https://static.igem.org/mediawiki/2021/7/79/T--Thessaly--ZagorinLogo.png)

![](https://static.igem.org/mediawiki/2021/8/8a/T--Thessaly--AnadrasiLogo.png)

![](https://static.igem.org/mediawiki/2021/7/7d/T--Thessaly--FlemingLogo.png)

![](https://static.igem.org/mediawiki/2021/3/3c/T--Thessaly--Lab_SolutionsLogo.png)

![](https://static.igem.org/mediawiki/2021/5/58/T--Thessaly--MediplanLogo.png)

![](https://static.igem.org/mediawiki/2021/9/94/T--Thessaly--MetrolabLogo.png)

![](https://static.igem.org/mediawiki/2021/f/f6/T--Thessaly--PindosLogo.png)

![](https://static.igem.org/mediawiki/2021/b/ba/T--Thessaly--SnapgeneLogo.png)

![](https://static.igem.org/mediawiki/2021/b/bf/T--Thessaly--SuperchipLogo.png)

![](https://static.igem.org/mediawiki/2021/0/0c/T--Thessaly--idt.png)

![](https://static.igem.org/mediawiki/2021/3/32/T--Thessaly--la-travel.png)

![](https://static.igem.org/mediawiki/2021/8/88/T--Thessaly--twist.png)