Notebook: Week 6
Preparations | Week 1 | Week 2 | Week 3 | Week 4 | Week 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Week 11 | Week 12 | Week 13 | Week 14 | Week 15 | Week 16
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Aims of the Week
Our main aim this week is to create a line of competent cells, by reinvigorating our previous cell line. This protocol began last week, and will be finished by Tuesday. At this point we can test the competency of the new line, and hopefully recommence our transformations. Also, with regards the wiki, we are hoping to make significant progress on its content. in particular we would like the get more of our information onto the Research page, and also set up a safety page, a protocols page, and a Lab Book. Also, Carina is working on some inforgraphics for our homepage, which will make the principles and aims of our projects much simpler. For the Rathenau debate, Erin hopes to make it half way through the first draft of the more detailed proposal Rathenau have asked for. Finally, for modelling, the aim is to develop the Matlab model for degradation, and we are waiting to hear back from Professor Rowley who will assist us with the degradation equation. In the meantime they will be working on buoyancy and the ocean model
Monday 16th July
Wet Lab - James and Aurelija continued the reinvigoration of the W3110 cell line today, after culturing it on agar over the weekend. Single colonies picked from minimal agar plates and used to seed 5ml cultures overnight in preparation for day 3 of protocol. We hope this will produce a more competent cell line, capable of more efficient transformation.
Modelling - Joanne, Aurelija and Erin continued their search for software for the ocean model representation, this included researching Ferret and Panoply, and lookinng into past iGEM modelling presentations. They are particularly impressed by Columbia 2011. Outcome: We have established the necessity to use Panoply software.
Website - Team Page Design - Carina has been working on the caricatures of the team for a 'meeting the team' image for the homepage.
Tuesday 17th July
Meeting - Wiki - Rhiannon met with Aurelija and Yeping to discuss the details for the Research page. The Research page is still a work in progress, and while we work on some diagrams to make our project more comprehensive, we want to ensure the text explains our project as clear as possible. Outcome: We have a draft overview for every module that explains each module in context of the others, and how the system works individually. We also hope to include a more detailed explanation of the decision making process.
Meeting - Degradation Module. Rhiannon and Yeping met to discuss the search for the laccase gene for the degradation module. Unfortunately, attempts to get hold of the C208 strain of Rhodococcus ruber has failed, which was our preferred choice of laccase gene. We researched other laccase genes which may have plastic degradation properties, as well as alternative enzymes. Outcome: It was very difficult to find a gene for which the sequence was available. We did find the sequence for one, but we are unsure of its plastic degradation properties.
Wet Lab - James and Aurelija completed their reinvigoration of the W3110 cell line, and Rhiannon transformed a sample of this cell line with a reference plasmid, to determine how successful the reinvigoration was. The transformed cells were placed in overnight culture.
Design - Carina has been working on the design for the homepage of the wiki. She is designing a series of images to explain our project 'in a nutshell', to enable visitors reading the more detailed area of the wiki to understand the context and scientific principles behind our project.
Wednesday 18th July
Wet Lab. - Leonard, James and Rhiannon tested the results of yesterdays transformation and found no colonies for our reinvigorated cells. This was concerning as it would take us another week to carry out a second reinvigoration – and so we decided to test the competency a second time. This time we would factor in the two different protocols we had used since commencing the project, and included our advisor Yanika’s cells, which are known to be competent. If one of the protocols worked for none of the cell types, we could consider the possibility that one protocol was less efficient than another. However, if it was the cell line that was incompetent, this too should be obvious.
Also Leonard and James were investigating ways to characterise the adhesive ability of Curli, and found a protocol that used water pressure between two discs to investigate shear force on Curlis. They have presented this to our supervisor Darren, and it will hopefully be possible to have the equipment made by the UCL workshop.
Modelling. Aurelija and Joanne continued working on degradation module. They managed to simulate first order enzyme kinetics on Excel . This is general degradation model that we now need to adopt to our system. Work done on degradation module today will lead to more complex degradation model that we will be specific to laccase. Once the model is finished it will be incorporated with ocean model to see the effect of degradation of laccase to plastic accumulation of micro-plastics.
Thursday 19th July
Wet Lab. We checked the results from yesterdays transformation and found that our first cell line was shown to be extremely competent – despite the test of its competency last week. This was great news, as it means we can progress with our project. Our second cell line, which had been reinvigorated, were found to be entirely incompetent – so we will not use these further. We also found that both protocols were usable, but the first protocol that we used was more successful.
Meeting – LMU Munich. Today we had meeting (via skype) with LMU team member Simon about characterisation and construction collaboration. We also discussed modelling collaboration possibilities. We decided on modelling workshop that we are going to prepare. The meeting with LMU Munich team is going to be next Thursday (last day of CAS conference). Plan is to prepare power point presentation which could concisely summarise the use of modelling in synthetic biology.
Meeting – Wet Lab. This meeting was organized by Darren Nesbeth our supervisor to discuss our progress. James, Leonard, Bouran and Aurelija updated the current situation regarding biobricks as well as new ways of characterising shear force of curli expression. With regards to competent cells, our supervisor encouraged us to use our competent cells from the first run, as they were shown to be highly competent’.
Poster. Aurelija has worked on the second text draft for Munich CAS conference poster. Carina worked on improving design for the poster, she created slightly different design for circuits from those in registry.
Meeting - Modelling. Today Erin and Aurelija had a meeting with Chris Barnes to discuss ABC sysbio software. We wanted to find out if this particular software would be useful for us. We decided to use this and the first model that we are going to be using it for is going to be degradation. It will be a great addition to our modelling arsenal (and also we might get to use the supercomputer!)
Meeting – Human Practice. The Human Practice team met with Darren to assess the progress of our Human Practice events. He suggested we contact more marine interest groups (Fishing for litter, CleanUp UK, Marine Society) and other societies, such as the Society of Biology to organise presentations and events with them. Darren has also contact the internation marine organisation (IMO) – an important marine oganisation – with the hope of organising a presentation with their committee. Finally, we are hoping to organise a beach clean-up at southbank.
Human Practice - Yeping was contacting galleries for an exhbition for the DIYbio event, and she found a place for the Speed Debating event.
Friday 20th July
Meeting - Modelling. Joanne, Aurelija and Yanika (one of our advisors) had a meeting with our supervisor Darren Nesbeth about our progress on different modules. We decided on having a rough draft of four models by the 1st of August. Our supervisor advised us to organise a meeting with Dr Dalby regarding the degradation module. Having a deadline will make us get our draft of the models done and then we will be able to build up on those.
Saturday 21st July
Poster. We are really excited! Today Carina, Aurelija and Bethan finished our first poster which summarises our project. As the poster is dedicated to Munich conference (to which six of us are going) we concentrated only on experimental and modelling plans. (photo with us and poster).
Sunday 22nd July
Presentation - Munich. Aurelija and Joanne finished the presentation for the LMU modelling meetup organised for the coming week. The aim is to organise a collaboration between the Munich iGEM team.
Reflections
This week has been a huge relief for the lab team, who had been growing increasingly concerned over the delay caused by the competency of our cells. However, now that we have found a protocol and a cell line that works, we are much relieved and keen to begin construction. Next week, we will be making up for the lost time, and driving the construction of our biobricks. While we have been delayed, we are still confident that all our aims are achievable, and we are looking forward to making progress. We also made a lot of progress with the content for the wiki - but this has not been published yet. This includes writing up a more comprehensive account of our research plans, and the designs for a more attractive and user-friendly homepage. Furthermore, the six of us that will be visiting Munich for the CAS conference in Synthetic Biology have been preparing all week to develop a collaboration with Munich, which has been very successful. Altogether, there has been a lot of pressure on this week but we have achieved all our aims.
Fail of the Week - Reinvigoration of Cell Line
Success of the Week - Original cell line proven more competent than we expected
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