From 2012.igem.org

Notebook: Week 4

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Aims For This Week

Our aim this week is to transform the Curli Cluster, Ribosome Binding Site and GFP biobrick for Module 1 – Binding to Plastics. The Modelling Crew are aiming to finish their ocean model, and begin modelling their 1st receptor model – for the Curli biobrick. The Human Practice team would also like to begin contacting other teams via the iGEM headquarters, to recruit for our radio show, and plan an exhibition at the Dana centre.

Monday 2nd July

Meeting – Construction/Characterisation Bouran, Leonard, James and Aurelija had a meeting with our main supervisor Darren Nesbeth to discuss our progress on construction/characterisation and to set realistic aims for the week. Several aspects of construction were discussed. The main focus was on the detection module which was based on the human oestrogen receptor (ER). From the data collected on ER folding and activity in prokaryotic cells, it became clear to us that it is extremely difficult to make ER function in prokaryotic cells. Consequently, we decided to look for alternatives for our detection system. However, as we already have ER on our kit we decided to evaluate its functionality soon to see if our assumptions about it were correct.

Modelling. Joanne, Aurelija and Erin started working on our first model for degradation module. First we looked for the right software. Initially we were considering both COPASI 4.8 as well as Matlab . We finally decided to use Matlab as it will give greater flexibility to our model. The aim for the degradation model is to represent degradation of polyethylene in two different environments: ocean as well as lab.

Human Practice Today Yeping, Martina and Carina planned our activities for UCL Open Day that will take place on Wednesday (4th of July). This is a great opportunity for us to highlight the problem of microplastics to society. We decided on the several interactive activities that we will be offering to the prospective UCL students during the open day. These will include a demonstration of motion video that has been created by our team members and which represents the idea behind our project; also we decided to prepare several questions about microplastics pollution to enhance the awareness amongst the perspective students. The interactive activity 'make your piece of art using plastic waste' was chosen as well; for this we will need to collect some plastic waste before Wednesday.

Today we researched posters from past iGEM teams in order to develop a clear understanding of what the main criteria are with regards to experimental details, visual representation as well as formatting. After looking through numerous examples, a list of 'dos and don'ts' was produced as well as the general structure of the poster. This gave us the basis for the poster format, as follows:

1. Abstract/Intro/Motivation
2. Modelling
3. Module 1 – detection
4. Module 2 – aggregation
5. Module 3 – degradation
6. Salinity, Buoyancy and Containment
7. Human practices /Public engagement
8. Conclusions/Achievements
9. Sponsors

Tuesday 3rd July

Wet lab - Agar PreparationToday our lab work was concerned with agar plate preparation which allow us to start transformations tomorrow. Three team member prepared the following agar plates: LB, LB+Ampicillin & LB+Chloramphenicol. This will allow us to begin biobrick transformation.

Modelling. We managed to deduce the equation for our degradation model that is based on specific activity of our enzyme of interest which is laccase. Tomorrow we will be able to write a code for this specific reaction on matlab. The equation deduced today is only initial, as we progress we might additional 'noise' representations to it.

Meeting - Wiki. Rhiannon organised a meeting with members of the construction crew to determine how to represent our research on the wiki. Outcome: We will provide users with a clear outline of how the modules interact, and a concise version of the design. We also want to provide a longer article which explains the process of decision-making regarding our choice of biobricks and methods. Finally, we decided to make our research pages accessible to those new to science by including a glossary of scientific terms

Achievement - Open day! Today Carina created an amazing poster that will be used during the open day tomorrow as our project representation! It shows all the land that will be available for all of us to inhabit once the plastic island is created.

Wednesday 4th July

Wet Lab - Transformation of Competent Cells. Todays lab team, Rhiannon, Bouran and Leonard, transformed cells with the Curli Cluster, Ribosome Binding Site, and GFP Biobrick. These three biobricks, which are involved in the adhesion module, were transformed using a standard heat shock protocol and incubated overnight. We have selected the adhesion module to start first, because we predict it will be the hardest to accomplish. We think we will have difficulties finding a receptor that E.coli will tolerate, and there is uncertain data on the success of the biobrick

Modelling. Erin invested time today learning how to use MATLAB GUI to make a user interface for our degradation module.Outcome: Erin produced a skeleton code which we can fit the equations into. This is signficant progress because this code can also be used to model our other modules

Human Practice - Outreach at UCL open day This was organised by Martina, with the purpose of reaching potential UCL students, and see what their opinion about our project is. Outcome: We became more proficient at presenting our research, and received some fresh input onto our project. This is important expeirience that will inform future events we organise.

Thursday 5th July

Wet Lab - PCR Reaction. Rhiannon, Yeping, Martina and Leonard were trained by Erick to set up a PCR reaction, use a PCR machine, and design primers. Also, upon checking the cells we transformed yesterday we were pleased to find that the Curli Cluster Biobrick and GFP Biobrick were successful. This demonstrated also that our work on making the cells competent was also successful. Colonies from these plates were subsequently picked and inoculated in LB medium and incubated overnight. Unfortunately the Ribosome Binding Site biobrick did not work, and so transformation will have to be repeated with a higher volume of competent cells.

Human Practice - Documentary. Documentary filming continued with the filming of several team members.

Friday 6th July

Wet Lab - PCR reaction. Yeping and Martina made a 1% Agarose gel to test the presence of our PCR product of PSB1C3 - a plasmid backbone we used in training yesterday. Also done was the miniprep of the plasmids containing Curli Cluster and GFP biobricks.

Meeting - Wiki. There has been some consideration of whether to include a 'lab book' alongside the notebook on the wiki. We have now decided to go forward with this idea. Some work will be done over the weekend to develop the clearest way to represent the research from different modules, and how we can make it sync with the notebook.

Poster. Today we had a meeting with Carina who is our adviser for visual representation. Previously produced structure outline and different types of visual representations of graphs and experimental modules were discussed. Outcome: We decided to create a more interconnected representation of our modules. As they really depend on each other, rigid separation of modules(boxes) was left out for now. First draft of the poster will be produced during the weekend.

Reflection

On reflection we have had a very successful week with regards to Human Practice and Modelling. Our open day was one of the key highlights - especially as it was the first display of Carina's plastic island poster. We have also had mounting success with out Crowdfunder campaign - an alliance between Human Practice and Sponsorship - which is sending our message out to other audiences and asking for their support. In the lab, we are still 'warming up' and training everyone, but it is our intention next week to throw ourselves into the research and get a lab book onto the wiki that demonstrates all of the successes and failures. In short, this week has been an exciting start to enacting our aims and next week will be much the same for the lab research.

Fail of the Week. Failure of the ribosome binding site biobrick to transform into bacteria.

Success of the Week: Our architect Carina's map of plastic island. Check out [http://www.plasticrepublic.org plasticrepublic.org]!

Team:University College London/Notebook/Week4