iGEM 2019 — Team Wageningen_UR

[![](https://2019.igem.org/wiki/images/8/8b/HQ_page_logo.jpg)](https://2019.igem.org)

# Team:Wageningen UR/Notebook

Xylencer

[![](https://static.igem.org/mediawiki/2019/thumb/6/6a/T--Wageningen_UR--Logo_only.png/370px-T--Wageningen_UR--Logo_only.png)XYLENCER](https://2019.igem.org/Team%3AWageningen_UR)
*menu*

* Project *arrow\_drop\_down*

+ [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
+ [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
+ [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)

* Results *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
+ [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
+ [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
+ [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)

* Notebook *arrow\_drop\_down*

+ [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
+ [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
+ [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)

* Safety *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
+ [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
+ [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
+ [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
+ [Safe By
  Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)

* Human Practices *arrow\_drop\_down*

+ [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
+ [Education &
  Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
+ [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
+ [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
+ [Voice of iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)

* About Us *arrow\_drop\_down*

+ [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
+ [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
+ [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
+ [About the Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

![Xylencer](https://static.igem.org/mediawiki/2019/thumb/4/48/T--Wageningen_UR--Logo-black.png/800px-T--Wageningen_UR--Logo-black.png)* + Project *arrow\_drop\_down*

    - [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
    - [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
    - [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)
  + Results *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
    - [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
    - [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
    - [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)
  + Notebook *arrow\_drop\_down*

    - [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
    - [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
    - [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)
  + Safety *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
    - [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
    - [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
    - [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
    - [Safe By
      Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)
  + Human Practices *arrow\_drop\_down*

    - [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
    - [Education &
      Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
    - [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
    - [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
    - [Voice of
      iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)
  + About Us *arrow\_drop\_down*

    - [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
    - [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
    - [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
    - [About the
      Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

# Labjournals

![Notebook](https://static.igem.org/mediawiki/2019/4/4f/T--Wageningen_UR--Notebook.jpg)

Below you will find the labjournal entries of the different team members. If you are interested in the weekly achievements or the struggles each of us went through, click the buttons below!

![subproject hexbadge](https://static.igem.org/mediawiki/2019/d/d0/T--Wageningen_UR--button_xylencer.png)

Team Notebook

In this notebook, you can see all the milestones of the complete Wageningen iGEM team.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Team)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/f/f1/T--Wageningen_UR--button_Alex.png)

Anti CRISPR proteins

Here you can find the journal of Alex. He worked on controlling the repression of the phage genome with Anti CRISPR proteins. This gives us more control over the phage repression.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Alex)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/2/2a/T--Wageningen_UR--button_Alba.png)

Molecular Timer Kill Switch

In the notebook of Alba you can read all about her time building a kill switch based on a molecular timer.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Alba)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/6/64/T--Wageningen_UR--button_Ben.png)

DSF Sensing Circuit

In Ben his labjournal, he detailed his journey to built a sensing circuit that could recognize *X. fastidiosa's* quorum sensing molecule DSF.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Ben)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/0/06/T--Wageningen_UR--button_Cleo.png)

Plant Immune Response

This is the notebook of Cleo. Here you can find more information on her journey from failed protein purification experiments to plant experiments. She worked on activating a plant immune response.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Cleo)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/b/bd/T--Wageningen_UR--button_Hetty.png)

Spatial Spread Model

Hetty modeled the spread of our Xylencer phage and  *X. fastidiosa*  to help us determine where and when to apply our therapy.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Hetty)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/3/38/T--Wageningen_UR--button_Marijn.png)

Cell-Free Expression System

Marijns notebook discusses not only Cell-Free Expression Systems, used for phage engineering and protein production, but also more general phage engineering approaches.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Marijn)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/7/77/T--Wageningen_UR--button_Niels.png)

Detection

This is Niels his notebook. Here you can read about how Niels researched our detection device.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Niels)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/4/43/T--Wageningen_UR--button_Robert.png)

Adhesion Protein Library

This is Roberts notebook. He built a protein library containing adhesion proteins that can bind to chitin.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Robert)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/c/c9/T--Wageningen_UR--button_Santi.png)

Yeast-based phage engineering

In Santi's notebook you can learn more about yeast-based phage engineering and other phage engineering methods to produce our Xylencer phage.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Santi)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/1/10/T--Wageningen_UR--button_bas.png)

Phage repression

This is Sebastiaans notebook, where you find information on the repression of phage genomes using both dead and active Cpf1.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Sebastiaan)

![subproject hexbadge](https://static.igem.org/mediawiki/2019/b/b8/T--Wageningen_UR--button_dennie.png)

Pathogenicity Prediction

This is the journal of Dennie. Here you will find more information on the prediction of non-pathogenic bacteria. The outcome of this subproject motivated the decision on what bacterium to use as our phage delivery bacterium.

[Read More*add*](https://2019.igem.org/Team%3AWageningen_UR/Notebook/Dennie)

##### Xylencer

A project by the Wageningen University & Research iGEM 2019 team.

##### Contact

*mail\_outline*
igemwageningen@gmail.com

##### Follow Us!

* [@igemwageningen](https://twitter.com/igemwageningen)
* [igem.wageningen](https://www.instagram.com/igem.wageningen/)
* [iGEMWageningen](https://www.facebook.com/pg/iGEMWageningen/posts/)
* [iGEM Wageningen](https://www.linkedin.com/in/igem-wageningen-333033191/)

Xylencer 2019

*swap\_vert*

* ###### SECTIONS
* [Back To Top](#home)


[![](https://2019.igem.org/wiki/images/8/8b/HQ_page_logo.jpg)](https://2019.igem.org)

# Team:Wageningen UR/Experiments

Xylencer

[![](https://static.igem.org/mediawiki/2019/thumb/6/6a/T--Wageningen_UR--Logo_only.png/370px-T--Wageningen_UR--Logo_only.png)XYLENCER](https://2019.igem.org/Team%3AWageningen_UR)
*menu*

* Project *arrow\_drop\_down*

+ [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
+ [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
+ [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)

* Results *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
+ [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
+ [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
+ [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)

* Notebook *arrow\_drop\_down*

+ [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
+ [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
+ [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)

* Safety *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
+ [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
+ [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
+ [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
+ [Safe By
  Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)

* Human Practices *arrow\_drop\_down*

+ [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
+ [Education &
  Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
+ [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
+ [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
+ [Voice of iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)

* About Us *arrow\_drop\_down*

+ [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
+ [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
+ [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
+ [About the Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

![Xylencer](https://static.igem.org/mediawiki/2019/thumb/4/48/T--Wageningen_UR--Logo-black.png/800px-T--Wageningen_UR--Logo-black.png)* + Project *arrow\_drop\_down*

    - [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
    - [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
    - [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)
  + Results *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
    - [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
    - [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
    - [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)
  + Notebook *arrow\_drop\_down*

    - [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
    - [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
    - [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)
  + Safety *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
    - [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
    - [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
    - [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
    - [Safe By
      Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)
  + Human Practices *arrow\_drop\_down*

    - [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
    - [Education &
      Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
    - [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
    - [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
    - [Voice of
      iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)
  + About Us *arrow\_drop\_down*

    - [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
    - [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
    - [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
    - [About the
      Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

# Protocols

![Group Photo](https://static.igem.org/mediawiki/2019/7/76/T--Wageningen_UR--protocols_header.jpg)

On this page, the protocols used by our team can be found. All protocols are uploaded on protocols.io.
Protocols are derived from iGEM, literature and suppliers or are designed by ourselves.

### Measurements Nucleotide and Protein Concentration

[Qubit](https://dx.doi.org/10.17504/protocols.io.e7sbhne%20)

[Nanodrop](https://dx.doi.org/10.17504/protocols.io.id2ca8e%20)

### Agarose gel

[Agarose Gel
Electrophoresis](https://dx.doi.org/10.17504/protocols.io.7ukhnuw%20)

[DNA
Purification
Agarose Gel](https://dx.doi.org/10.17504/protocols.io.7hrhj56)

### PCR

[Overlap Extension
PCR](https://dx.doi.org/10.17504/protocols.io.psndnde%20)

[OneTaq](https://dx.doi.org/10.17504/protocols.io.cq8vzv%20)

[Phusion](https://dx.doi.org/10.17504/protocols.io.irrcd56%20%20)

[Phire](https://dx.doi.org/10.17504/protocols.io.7kkhkuw%20)

[Q5](https://dx.doi.org/10.17504/protocols.io.cidua5%20)

### Cloning

[Chemically
Competent Cells](https://dx.doi.org/10.17504/protocols.io.7jwhkpe)

[Heat Shock
Transformation](https://dx.doi.org/10.17504/protocols.io.isxcefn%20)

[Electrocompetent
cells](https://dx.doi.org/10.17504/protocols.io.7jrhkm6%20)

[Electroporation](https://dx.doi.org/10.17504/protocols.io.crgv3v%20)

[Mix&Go
Competent Cells](https://dx.doi.org/10.17504/protocols.io.7jnhkme)

[Ligation](https://dx.doi.org/10.17504/protocols.io.irscd6e%20)

[Restriction
Enzyme
Digestion](https://dx.doi.org/10.17504/protocols.io.isycefw%20)

[Gibson](https://dx.doi.org/10.17504/protocols.io.cdms45%20%20)

[Golden Gate
Assembly](https://dx.doi.org/10.17504/protocols.io.7kfhktn)

[Removal
Kanamycin-R
gene](https://dx.doi.org/10.17504/protocols.io.7jqhkmw%20)

### RNA Isolation

[RNeasy](https://dx.doi.org/10.17504/protocols.io.ssbeean%20)

[Trizol](https://dx.doi.org/10.17504/protocols.io.zvwf67e%20%20)

[DNase I
Treatment](https://dx.doi.org/10.17504/protocols.io.75ghq3w%20%20%20)

### Protein Expression and Purification

[Protein
Expression](https://dx.doi.org/10.17504/protocols.io.7i4hkgw%20)

[Protein
Sample Preparation](https://dx.doi.org/10.17504/protocols.io.7p9hmr6%20)

[Purification
Gravity-Column](https://dx.doi.org/10.17504/protocols.io.7qahmse%20)

[Purification
FPLC](https://dx.doi.org/10.17504/protocols.io.7qchmsw%20%20)

[SDS Page
Gel](https://dx.doi.org/10.17504/protocols.io.7syhnfw%20)

[Tricine SDS
Page Gel](https://dx.doi.org/10.17504/protocols.io.7sxhnfn%20%20)

### Chitin Binding

[Chitin Binding &
Bradford
Assay](https://dx.doi.org/10.17504/protocols.io.8akhscw%20%20)

[Chitin Magnetic
Beads](https://Dx.doi.org/10.17504/protocols.io.8anhsde%20%20%20%20)

### Phages

[Phage Stock](https://dx.doi.org/10.17504/protocols.io.7kthkwn%20)

[Phage
Genome Isolation](https://dx.doi.org/10.17504/protocols.io.7kvhkw6)

[Spot Assay](https://dx.doi.org/10.17504/protocols.io.7i9hkh6%20)

[Plack Assay](https://dx.doi.org/10.17504/protocols.io.7i8hkhw%20%20)

[Genome
Concatermerization](https://dx.doi.org/10.17504/protocols.io.7k3hkyn%20)

[TXTL Phage
Production](https://dx.doi.org/10.17504/protocols.io.77thrnn%20%20)

### Phage Repression

[Growth Curve
Analysis](https://dx.doi.org/10.17504/protocols.io.77jhrkn%20)

[GFP-RFP Plate
Reader Assay](https://dx.doi.org/10.17504/protocols.io.784hryw%20)

[Fluorescent Loss
Assay Acr](https://dx.doi.org/10.17504/protocols.io.7mfhk3n)

### Yeast

[Yeast Competent Cells](https://dx.doi.org/10.17504/protocols.io.7jphkmn%20)

[Transformation](https://dx.doi.org/10.17504/protocols.io.7jkhkkw)

### Molecular Timer

[72h
OD&GFP
measurements](https://dx.doi.org/10.17504/protocols.io.8aihsce)

### Plants

[Plant
Infection](https://dx.doi.org/10.17504/protocols.io.725hqg6%20)

[Xylem
Inoculation](https://dx.doi.org/10.17504/protocols.io.id2ca8e%20)

[Lesion
Size](https://dx.doi.org/10.17504/protocols.io.8bshsne)

### Insects

[Capture](https://dx.doi.org/10.17504/protocols.io.8jdhui6)

[Storage](https://dx.doi.org/10.17504/protocols.io.8jfhujn)

[Transmission
Assay](https://dx.doi.org/10.17504/protocols.io.8jghujw)

### LAMP

[LAMP assay](https://dx.doi.org/10.17504/protocols.io.8jahuie%20)

[LAMP
Paraffin encasing](https://dx.doi.org/10.17504/protocols.io.8jbhuin%20)

[*X.
fastidiosa* extraction](https://dx.doi.org/10.17504/protocols.io.8i8huhw)

![divider](https://static.igem.org/mediawiki/2019/3/3c/T--Wageningen_UR--leaf_divider_orange.png)

##### Xylencer

A project by the Wageningen University & Research iGEM 2019 team.

##### Contact

*mail\_outline*
igemwageningen@gmail.com

##### Follow Us!

* [@igemwageningen](https://twitter.com/igemwageningen)
* [igem.wageningen](https://www.instagram.com/igem.wageningen/)
* [iGEMWageningen](https://www.facebook.com/pg/iGEMWageningen/posts/)
* [iGEM Wageningen](https://www.linkedin.com/in/igem-wageningen-333033191/)

Xylencer 2019

*swap\_vert*

* ###### SECTIONS
* [Back To Top](#home)

[![](https://2019.igem.org/wiki/images/8/8b/HQ_page_logo.jpg)](https://2019.igem.org)

# Team:Wageningen UR/Results

Xylencer

[![](https://static.igem.org/mediawiki/2019/thumb/6/6a/T--Wageningen_UR--Logo_only.png/370px-T--Wageningen_UR--Logo_only.png)XYLENCER](https://2019.igem.org/Team%3AWageningen_UR)
*menu*

* Project *arrow\_drop\_down*

+ [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
+ [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
+ [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)

* Results *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
+ [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
+ [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
+ [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)

* Notebook *arrow\_drop\_down*

+ [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
+ [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
+ [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)

* Safety *arrow\_drop\_down*

+ [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
+ [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
+ [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
+ [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
+ [Safe By
  Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)

* Human Practices *arrow\_drop\_down*

+ [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
+ [Education &
  Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
+ [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
+ [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
+ [Voice of iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)

* About Us *arrow\_drop\_down*

+ [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
+ [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
+ [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
+ [About the Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

![Xylencer](https://static.igem.org/mediawiki/2019/thumb/4/48/T--Wageningen_UR--Logo-black.png/800px-T--Wageningen_UR--Logo-black.png)* + Project *arrow\_drop\_down*

    - [Description](https://2019.igem.org/Team%3AWageningen_UR/Description)
    - [Design](https://2019.igem.org/Team%3AWageningen_UR/Design)
    - [Background](https://2019.igem.org/Team%3AWageningen_UR/Background)
  + Results *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Results)
    - [Modeling](https://2019.igem.org/Team%3AWageningen_UR/Model)
    - [Demonstrate](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate)
    - [Achievements](https://2019.igem.org/Team%3AWageningen_UR/Achievements)
  + Notebook *arrow\_drop\_down*

    - [Labjournals](https://2019.igem.org/Team%3AWageningen_UR/Notebook)
    - [Protocols](https://2019.igem.org/Team%3AWageningen_UR/Experiments)
    - [Parts](https://2019.igem.org/Team%3AWageningen_UR/Parts)
  + Safety *arrow\_drop\_down*

    - [Overview](https://2019.igem.org/Team%3AWageningen_UR/Safety)
    - [Biosafety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosafety)
    - [Biosecurity](https://2019.igem.org/Team%3AWageningen_UR/Safety/Biosecurity)
    - [Lab Safety](https://2019.igem.org/Team%3AWageningen_UR/Safety/Lab_Safety)
    - [Safe By
      Design](https://2019.igem.org/Team%3AWageningen_UR/Safety/Safe_By_Design)
  + Human Practices *arrow\_drop\_down*

    - [Human Practices](https://2019.igem.org/Team%3AWageningen_UR/Human_Practices)
    - [Education &
      Engagement](https://2019.igem.org/Team%3AWageningen_UR/Public_Engagement)
    - [Collaborations](https://2019.igem.org/Team%3AWageningen_UR/Collaborations)
    - [Ethics](https://2019.igem.org/Team%3AWageningen_UR/Ethics)
    - [Voice of
      iGEM](https://2019.igem.org/Team%3AWageningen_UR/Voice)
  + About Us *arrow\_drop\_down*

    - [The Team](https://2019.igem.org/Team%3AWageningen_UR/Team)
    - [Attributions](https://2019.igem.org/Team%3AWageningen_UR/Attributions)
    - [Sponsors](https://2019.igem.org/Team%3AWageningen_UR/Sponsors)
    - [About the
      Wiki](https://2019.igem.org/Team%3AWageningen_UR/About)

# Results

![Results Header](https://static.igem.org/mediawiki/2019/8/8e/T--Wageningen_UR--Leaf_zoom.jpg)

On this page, we proudly present an birds-eye view of the results of the entire Xylencer project. Here you
will find a summary of the most important conclusions drawn from each of the lines of research within
Xylencer. In-depth reports on each topic can be found by clicking on their corresponding buttons below. For
a more extensive report on our scientific highlights, including supporting figures, visit our [demonstrate page](https://2019.igem.org/Team%3AWageningen_UR/Demonstrate).

### Detection

The first pillar of Xylencer is focused on detecting *X.
fastidiosa*
by an automated tool enabling in-field testing based on Loop-mediated isothermal
amplification (LAMP). Proof of principles for the key mechanisms of this device was obtained
through multiple tests. We showed that we could perform LAMP on bacteria with *X.
fastidiosa*
specific primers. To ensure LAMP reagents stability over time and temperature influences,
and to prevent insects from ingesting it, we showed that the reaction mixture can be
preserved in paraffin at a slight reduction in short-term efficiency. By performing
insect-transmission assays, we were able to show that insects fed on a plant mimic and could
transfer bacteria from a source to this plant mimic. We determined the optimal feeding
medium concentration, namely 0.2% sucrose, that still allows LAMP to function. Finally, in
collaboration with The Netherlands food and consumer product safety authority (NVWA), we conducted a LAMP test on leaf extracts
from infected plants. These tests demonstrated that a LAMP reaction temperature of 65 °C can
lyse the bacterial cell wall and make their DNA accessible for the LAMP primers,
successfully detecting X. fastidiosa with a visible readout.

[![Detection Device](https://static.igem.org/mediawiki/2019/7/77/T--Wageningen_UR--button_Niels.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Detection_Device)

###### Detection Device

![](https://static.igem.org/mediawiki/2019/d/d3/T--Wageningen_UR--Detection_pillar.png)

Overview of the detection pillar: Here you can see how the plant mimic attracts
an
insect, detects *X. fastidiosa*, and shows a visible color change to alert the
user.

### Delivery

In the second pillar of Xylencer, we revolutionized phage delivery with
the Phage Delivery Bacterium (PDB). To ensure our PDB would be safe, we trained a machine
learning model on a manually curated pathogenicity dataset and established a genetic basis for
non-pathogenicity. This allowed us to select *Xanthomonas arboricola* CITA 44 as our PDB.
To
direct the release of the pages to the areas of infection, we designed a circuit that controlled
phage expression based on *X. fastidiosa*’s presence. For this circuit, we created a
chimeric *X.
fastidiosa* quorum-sensing receptor, linked to a tightly controlled riboswitch. This
riboswitch
is coupled to a CRISPR-dCas circuit that is designed to control phage expression. We showed that
this system can control the expression of the phage operon. Besides this, we showed that dCas
repression can be controlled with an Anti-CRISPR system, allowing us to fine-tune the circuit.
To contain the spread of our PDB and guarantee biosafety, we explored the possibility of a timed
kill-switch, that would only prove lethal after a set amount of time. We developed a circuit
that increases protein expression over a period of three days. This system can be linked to a
toxin/anti-toxin system to achieve delayed induced lethality.

[![Pathogenicity Analysis](https://static.igem.org/mediawiki/2019/b/b8/T--Wageningen_UR--button_dennie.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Pathogenicity)

###### Pathogenicity analysis

[![Sensing Circuit](https://static.igem.org/mediawiki/2019/6/64/T--Wageningen_UR--button_Ben.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/DSF_Sensing_Circuit)

###### Sensing Circuit

[![Phage Repression](https://static.igem.org/mediawiki/2019/f/f1/T--Wageningen_UR--button_Alex.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Phage_Repression)

###### Phage Repression

[![Kill Switch](https://static.igem.org/mediawiki/2019/2/2a/T--Wageningen_UR--button_Alba.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Kill_Switch)

###### Kill Switch

![](https://static.igem.org/mediawiki/2019/5/50/T--Wageningen_UR--Delivery_pillar.png)

Overview of the delivery pillar. Top-left: *X. fastidiosa* (purple bacterium)
secretes
a quorum sensing molecule (green). Top-right: The quorum molecule is detected by the phage
delivery bacterium (green-bacterium). Bottom-right: Detections of the quorum sensing
molecule
triggers production and release of the phages, it also triggers the release of PAMPS (orange
compound, described in remediation). Bottom-left: The released phages infect *X.
fastidiosa*
cells and lyse them, releasing even more phages.

#### Drag the Button to Reveal the PDB

![PDB Closed](https://static.igem.org/mediawiki/2019/d/dd/T--Wageningen_UR--PDB_closed.png)

![PDB Opened](https://static.igem.org/mediawiki/2019/8/8a/T--Wageningen_UR--PDB_open.png)

*An overview of the phage delivery bacterium. Left to right: The* X. fastidiosa *quorum
sensing
molecule (green triangles) is detected by the chimeric receptor, triggering a signaling cascade.
This
signaling cascade switches the riboswitch on allowing for expression of anti-CRISPR. The anti-CRISPR
then relieves repression of the phage genome, allowing the Xylencer phages to be produced,
eventually
lysing the cell.*

### Remediation

In the third pillar, remediation, we improved phage therapy by forging an alliance with the
plant. This is achieved by alerting the plant to the presence of the pathogen, through
artificially triggering a systemic plant immune response. To this end, Xylencer phages
encode for a Pathogen Associated Molecular Pattern (PAMP): flg22. Phage infection and lysis of
*X. fastidiosa* would set these PAMPs free within the plant’s xylem and trigger plant
immune
responses. We performed plant experiments by infecting *Brassica oleacea* with a
close
relative of *X. fastidiosa*, *Xanthomonas campestris* pv. Campestris and
showed that inoculation
of flg22 into the plant’s xylem decreases disease symptoms during the crucial early stages
of infection.

[![Plant Immune Response](https://static.igem.org/mediawiki/2019/0/06/T--Wageningen_UR--button_Cleo.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Plant_Immune_Response)

###### Plant Immune Response

![](https://static.igem.org/mediawiki/2019/e/ec/T--Wageningen_UR--Remediation_pillar.png)

Activation of the plant's immune system helping to fend of *X.
fastidiosa*.

### Spread

The final pillar includes the design of a self-spreading mechanism for
the Xylencer phages. This allows our therapy to reach infected plants that would otherwise be
missed by conventional methods. In order to achieve this, Xylencer phages will be bound to the
mouthparts of the insects that spread *X. fastidiosa* by fusing adhesion proteins to their capsid.
We demonstrated the ability of *X. fastidiosa* derived adhesion protein PD1764 to bind to
structures found in the insect's mouthparts. Furthermore, we showed that we can produce a mix of
wild-type and adhesion capsid fusion proteins, allowing for increased stability of the phage capsid
by only partially saturating it with fusion proteins. Using cell-free transcription-translation
we produced *E. coli* phage T7 and phage Lambda capsid proteins, serving as a model for the
Xylencer phages. We constructed a spatial spread model that can simulate the effectivity and
spread of the Xylencer phages. This model showed that our Xylencer phages would be effective at
combatting *X. fastidiosa*, surpassing current methods.

[![Adhesion Library](https://static.igem.org/mediawiki/2019/4/43/T--Wageningen_UR--button_Robert.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Adhesion_Library)

###### Adhesion Library

[![Phage Engineering](https://static.igem.org/mediawiki/2019/d/d4/T--Wageningen_UR--phage_engineering.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Phage_Engineering)

###### Phage Engineering

[![Phage Engineering](https://static.igem.org/mediawiki/2019/9/93/T--Wageningen_UR--alternative_phage_engineering.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Alternative_Phage_Engineering)

###### Alternative Phage Engineering

[![Spatial Spread Model](https://static.igem.org/mediawiki/2019/b/bd/T--Wageningen_UR--button_Hetty.png)](https://2019.igem.org/Team%3AWageningen_UR/Results/Spatial_Spread)

###### Spatial Spread Model

![](https://static.igem.org/mediawiki/2019/c/ce/T--Wageningen_UR--Spread_pillar.png)

Overview of the spread pillar. Bottom-right: The insect vector feeds on a plant that
has been cured by the Xylencer therapy, this plant contains the Xylencer phage. Bottom-left:
The Xylencer phages attach to the stylet of the insect using their adhesion protein.
Top-left: The insect then spreads the Xylencer phages to an untreated field infected with
*X.
fastidiosa* by feeding on these trees. Top-right: The Xylencer phages cure the
diseased
plants of their infection.

##### Xylencer

A project by the Wageningen University & Research iGEM 2019 team.

##### Contact

*mail\_outline*
igemwageningen@gmail.com

##### Follow Us!

* [@igemwageningen](https://twitter.com/igemwageningen)
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Xylencer 2019

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