Difference between revisions of "Team:Exeter/Log"

March 2^nd

Log entry

We had our first meet up today. To get to know eachother better in small teams we were set the task to construct a spaghetti tower to hold a marshmallow as high as possible, only a bit of tape and some string.

We met the 2015 Exeter team who gave us advice concerning the iGEM competition.

March 9^th

Log entry

Today Professor John Love gave us an introductory synthetic biology lecture which helpful to those of us who are not studying biosciences.

March 16^th

Log entry

We had our team and individual photos taken today by Aaron Yeung of Exeter Photo Society. This was followed up by our first group brainstorm session.

ideas img

The preliminary ideas include:

Finding traces of peanut
Degrading plastic
Creating rocket fuel
Soft bioluminescent lighting
Making a screen using bioluminescence
Making plastic
Detecting STD's

We then split up into groups to research broad categories, including food security, diseases and space, and plan to feedback to each other in the next meeting.

March 18^th

Log entry

You can see below the team photo taken the other day.

team img

March 22^nd

Log entry

Today we were given a talk by Markus Gershater, the cofounder of Synthace

twitter img

We discussed the foundations of synthetic biology itself, the core problems in the synthetic biology field and potential routes to go down with our project. Our following discussion focused on taking a measurements based approach to iGEM instead, testing components to identify their optimum operating conditions. The idea was to attempt to identify how environmentally dependant the biobricks are.

March 23^rd

Log entry

Today, we had a short discussion about some of the points discussed on 16.03.16 with our iGEM advisors to decide which were viable routes to pursue, and which would likely not work or take far too long.

We also had a tour around the main biology lab, and were introduced to the equipment and machinery we will be using.

May 23^rd

Log entry

Today we discussed the medal criteria and outreach ideas.

These ideas include talking with experts in the field, holding introductory workshops in schools to introduce children to synthetic biology and running stalls at science fairs.

Potential ideas:

Synthesizing a microportha virus
Kill switches evolutionary stability, potential to oscillate a kill switch (genetic clocks)
Synthesizing PAF with E.Coli and combating fungal diseases in plants

May 24^th

Log entry

We mostly focused on how to develop a good grounding for our human practises. We want to educate people about synthetic biology. We also set up a YouTube channel to allow us to start uploading videos.

May 25^th

Log entry

We have decided to do daily vlog to recap our progress throughout the day.

We discussed the kill switch idea more in depth, looking at the possibility of using a lab grown amino acid to trigger the kill switch.

repressillator img

Above an idea relating PAF and repressilators can be seen.

We identified resources that we could offer for our collaborations. Our main specialties include: modelling and data processing, some specialised equipment and mentoring new teams.

We also began to identify key academics and companies we can establish connections with to better inform our project, and develop a deeper understanding of the synthetic biology involved in kill switches, antifungal proteins and repressilators.

At this point in time, we have been in contact with teams based in: Leiden, Newcastle, Cardiff and Costa Rica. We are hoping to set up Skype meeting with them in the near future.

May 26^th

Log entry

Today we looked at Astro Synbio, an educational web series about synthetic biology and space, and developed the idea of creating an episode of our own that could include developing rocket fuel, terraforming, soil purification (allowing a collaboration with Leiden), water purification, crop growth and food in space. For the rest of the day we started creating the iGEM wiki and learning basic HTML.

May 27^th

Log entry

We discussed the idea of a killswitch in the pathogen responsible for Rice Blast disease and realised that we would need to spray an entire rice field for it to be effective. To overcome this issue we came up with the idea of using a genetic clock to construct a “time bomb kill switch” in which the killswitch was triggered by the build up of a specific protein.

We are very excited about the idea of creating a board game targeted at secondary school children. Some potential concepts included a set cards that have basic questions, a build-a-cell game where the first to obtain all necessary parts wins or a combination of the two.

May 30^th

Log entry

Today was mostly focused on writing up a Penicillium antifungal protein (PAF) proposal for the Synenergene grant and expanding an idea of creating a CRISPR/Cas9 killswitch. Unfortunately there much more we can do at the moment beyond fleshing out ideas and emailing academics.

May 31^st

Log entry

It was a very exciting day as today we recorded our very first blog entry! Vlogs are a great way for both the public and any absent iGEM members to keep up with our project and see what progress we have made in bite sized chunks. At the moment we have decided to publish them on YouTube two weeks after filming to allow time for any editing.

We opened dialogue with teams based in Newcastle and Costa Rica today and have scheduled a Skype meeting with Leiden for tomorrow.

A few of the team members met with Professor Rob Beardmore to discuss our current ideas. He gave us some great feedback on potential flaws which we are now working on fixing.

June 1^st

Log entry

To start off the day today we had a Skype call with the Leiden iGEM team where we discussed potential ideas and what we’re currently looking at doing for human practises.

There was a suggestion to combine the PAF and the killswitch idea to create a system with a reliable and well characterised killswitch that could be used outside the laboratory in the environment.

June 2^nd

Log entry

We met with Dr Paul James to discuss the project and the human practises ideas.

Some members of the team met with Professor Peter Winlove who raised the following questions in regards for our PAF idea:

How much protein do you need to kill rice blast?
Could we model how much protein is required to kill rice blast?
Could we have a timer system as a killswitch?
Is there a heat change associated with fungal disease?
Could we use this heat change to turn on PAF production?
Could we engineer a bug to manufacture particles of different magnetic properties?
Can we change the structure of flagella on E. coli to change their swimming patterns/motions?
What is our end point/scenario? How can we work back from there?

June 3^rd

Log entry

Today we had a meeting with Dr Nick Harmer who suggested we look at secretion systems in regards to the PAF idea. This would allow PAF to pass through the outer membrane of E.coli. He also put forward the idea of using a gram positive bacteria to get around the problem of the double membrane of the gram-negative E. coli.

There was a new idea created involving a novel method of spreading a killswitch throughout a population of fungi: a virus would act as a vector for a killswitch which would pass throughout a population of fungi via its natural sexual reproduction.. The virus could have the genes that codes for the production of PAF so when the viral genetic information is inserted into the fungus PAF would be expressed, killing the fungus. However our main concern thus far is how the public would perceive an idea like this.

June 6^th

Log entry

There was a new idea created involving a novel method of spreading a killswitch throughout a population of fungi: a virus would act as a vector for a killswitch which would pass throughout a population of fungi via its natural sexual reproduction.. The virus could have the genes that codes for the production of PAF so when the viral genetic information is inserted into the fungus PAF would be expressed, killing the fungus. However our main concern thus far is how the public would perceive an idea like this.

Several emails were sent out today to various other iGEM teams, companies and people, including Adam Zivanic from the City of London school for potential human practises collaboration with his iGEM team, London Biohackspace; EMW Bookshop Community Lab, Street Bio, Genspace and William and Mary.

June 7^th

Log entry

We established contact today with Adam Zivanic today, head of Biology at City of London School. They’re entering a high school team into iGEM this year and the team seemed very interested in collaborating with us on the board game front. We intend to meet with them when we travel to London in August.

Following this we carried out research into current education on synthetic biology and unfortunately found there was very little education prior to postgraduate level, despite there being a fair amount of funding going into the field. In 2015, £40M was invested into synthetic biology research in the UK (see link) ¹. This has encouraged our team to want to look at potential ways to close that gap between funding and education, and introduce the synthetic biology earlier on. One way of doing this is developing a new module for the university.

A few members of the team worked through papers on the killswitch KillerRed to gauge the current level of characterisation for it.

http://www.bbsrc.ac.uk/news/policy/2015/150129-pr-business-secretary-40m-for-synbio/

June 8^th

Log entry

The work experience student worked through iGEM Labster and his feedback was that he thinks it needs better prompts to be more accessible. We spoke with him about his current GCSE syllabus, and found that synthetic biology could be taught alongside genetic modification and cloning. However, some students are confused between the differences of cloning and genetic modification. He also gave us his thoughts on the board game and overall he said it looks fun.

Peter Reader, a past Exeter iGEM team member, spoke with us about potential problems to be careful of. He advised us to be wary of parts which haven’t been used much, as it tends to mean they haven’t been characterised well meaning more room for error. He also advised meeting multiple lecturers to have them pull our ideas apart, point out potential flaws and make suggestions to amend these problems.

We also spoke with Ryan Edginton, a former iGEM member, about the wiki and the presentation. He gave us basic facts about the wiki, for example what happens on the day of the wiki freeze and required content.

He also gave us some more detailed advice about developing a brand for the project, including a logo and colour scheme, establishing a team leader for each section, fully defining the meaning of all equations used and explain the symbols and constants involved, and constantly keep testing on all internet browsers to ensure the wiki works on all browsers.

June 9^th

Log entry

A lot of today was spent working through past team’s projects to see what could be relevant to our project ideas, what couldn’t be completed in the past, what will be difficult, and what we could expand on for our own idea. There was a suggestion of using a delivery system, or developing a way to stick E. coli to the fungal cell wall to ensure delivery of PAF is direct.

We got our first card prototypes for the board game printed off today which we were all excited about. As shown below, there are four card types: promoters, ribosome binding sites, protein coding regions and terminators. We intend to make some rare and super rare cards.

Get image working! Prototype cards

In addition to this, we started developing an instruction manual to accompany the game.

We are working on how to “brand” the project by looking at different colour schemes so that when we start coding the wiki we have a general idea of what we want it to look like. Alan has put together a powerpoint introducing basic HTML concepts, so that the team members who haven’t coded before can learn.

June 10^th

Log entry

We made some slight changes to the board game cards today so we can incorporate the concept of scenarios, where the cards played by a team have to be justified. This makes the game a lot more interactive and encourages students to actually think about what cards they play, as opposed to simply playing cards to earn points.

There was a lot of looking at past wikis so we could start forming an idea of what we wanted the layout of our wiki to look like.

We discussed the PAF further. We did some research into the use of a secretion system to get PAF out of E. coli and noted how the mode of action of PAF is currently not well understood. We looked at how our project can tackle this problem, for example attatching GFP to the signal peptide known on PAF would allow us to “follow” it.

It was the work experience student’s last day with us today so we him what he had learned about synthetic biology during the week, how his perceptions towards the topic had changed and if he had enjoyed the week.

June 13^th

Log entry

Today was the first day with the majority of the team back together, so the key focus today was sitting down and re-brainstorming ideas before we settle on one. As can be seen in the mind map below, the PAF idea was fleshed out and developed in more detail.

We wrote an improved proposal for both of the measurement projects about killswitches and genome integration.

get image working! Brainstorm

Some of the team spoke to Dr Mark Ramsdale, a fungal cell biologist. He gave feedback on the PAF dea, raising the question as to why we wanted to express PAF in E. coli as opposed to a fungus. Dr Ramsdale pointed out that due to how small PAF was as a protein, using GFP as a tag could affect how it is transported around the cell, and therefore skew the results. He told us about fungi specific promoters in Neurospora crassa which acts as a Circadian Clock using light and heat as inducers.

This prompted a discussion about using a viral vector that contains a killswitch that. The viral vector will insert its genetic information in the host, a strain of Magnaportha oryzae. The Magnaportha oryzae would breed and the population would be infected with the virus containing the killswitch. The possibility of this being a heat activated killswitch was considered, so during the warmer months when plants grow the killswitch fungi is active to produce PAF.

Some of the team also met with Dr Nicky King, who is going to get us a stand at the Big Bang Science Fair for the South West (29.06.16) and the Britain Needs Scientists fair (06.07.16). She also discussed with us the possibility of writing an undergraduate Synthetic Biology module to be taken up by either Natural Sciences or Biosciences, and seemed very keen on the possibility of this.

We also met with Professor Rob Beardmore to discuss the genome integration idea. He said it sounds interesting but raised concerns about if it would even be possible to mathematically model it. He also liked the killswitch idea and said testing KillerRed and KillerOrange under a gradient of slowly increasing light intensity could be the strongest selector for resistance.

Get image working! Brainstorm

We also experimented with the vlog format, testing out the dynamics of using three people instead of two which we felt it worked a lot better.

June 14^th

Log entry

Some of the team skyped with the Newcastle iGEM team. We discussed potential for collaboration – they offered the skills of their computer scientists to help us with coding, and we offered the skills of our physicists to help with their understanding of electronics. We decided to Skype again in future and keep in touch with each other, as it seems there’s a lot of potential for collaborating.

We also spoke with Alice Mills, a Physics outreach officer, who is helping us develop ideas for the Physics/Maths/Computing side of our Synthetic Biology module. She is also going to put us in touch with a few different schools to allow us to test our board game with a range of classes.

June 15^th

Log entry

We made major progress with the board game today. Everyone put forward ideas to help improve it. The preliminary designs have been made and have been test printed. The ATCG counters used to keep score have been 3D printed in plastic.

We also had several meeting today. Dr Harmer spoke with us about the potential pitfalls of the PAF idea and how we can get around/solve them. Professor Smirnoff spoke with us about several of our ideas, giving his opinion of what he thought will work best.

We started talking with the iGEM teams based in Westminster, Purdue and UNL with the intention of setting up collaborations with them.

June 16^th

Log entry

Today was a day focused heavily on developing our human practises. We started constructing a booklet of rules and brainstormed ideas for the scenario cards. The role of scenario cards is to make the players justify their use of Promoter cards and how well it fits the scenario dictates how many points the card scores.

We met with two of our instructors today. Jamie Gilman discussed the benefits of using design of experiments with us. Dr Chloe Milner discussed our current potential ideas with us and ordered a gBlock for us so we could test whether or not PAF could be expressed in E. coli, which is one of the main concerns we have before we commit to a project.

June 17^th

Log entry

Everyone in the team had a lab induction today with Dagmara Kolak to familiarise ourselves with the the Mezzanine lab.

We split into three teams to research the three main ideas and prepare to present them to academics from various disciplines later on:

PAF (Pablo, Alice, Andy and Hannah)
Viral Vector Killswitch (Jack, Eloise and Leanne)
Measurement- kill switches and genome integration (Dan, Emily, Joel and Alan)

We hope to get expert feedback on flaws in our projects and what’s actually possible to complete within the summer.

June 18^th

Log entry

The board game was the focus of today again as we are trying to finish the first draft in time for the Judd school visit and continuing working on ideas and presentations in anticipation for the meeting with academics.

June 19^th

Log entry

We met with Dr Burton today who helped us a lot with ideas on the human practises side. She gave us contacts to help with trying to get synthetic biology on the GCSE syllabus, told us about a microbiology grant we were eligible for and gave us the idea of doing a “walkthrough science” video in collaboration with Soapbox Science.

We also started talking with Grace Fleet over email, who is a public engagement officer, focusing on scientific research. We intend to set up a meeting with her in the coming days.

June 20^th

Log entry

Part of the team went to the lab today to go through the Interlab study materials that we had been sent, but we couldn’t start as our kit was missing parts. We have emailed iGEM headquarters in the hopes they can be quickly replaced.

We also began working on the synthetic biology module. We currently have 7 potential textbooks which we’re taking notes on to inform our decision on what could and should go into the module, and which textbooks will best supplement this. We ensured that the textbooks were available online through the university, making them accessible to all who wanted to take the module.

The potential textbooks include:

The Analysis of Biological Data by Whitlock and Schulter
Understanding Bioinformatics by Zvelebil and Baum
Core Maths for the Biosciences by Reed
Next Generation DNA Sequencing Informatics by Brown
Molecular Biology of the Gene by Watson, Baker, Bell, Gann, Levine and Losick
Choosing and Using Statistics: A Biologist’s Guide by Dytham
Biostatistical Analysis by Zar

June 21^st

Log entry

Today was the first day of the Judd School trip. Alice, Andy, Alan and Jack left early this morning to travel to the school, and will be focusing on running board game sessions with the students, filming the game being played and filming interviews with the teachers and students.

The rest of the team remained at the university and continued to work through textbooks for the synthetic biology module.

We officially signed up for the interlab study today as and today’s lab team for the day attempted to test what we had with some competent cells prepared by Jamie Gilman to see what we could accomplish with the current kit.

Curves on the plate reader in the lab to prepare for measuring fluorescence levels, and incubated some flask cultures overnight in the 37 shaking incubator.

We also began to discuss possible activities we could include on our stand at the upcoming Big Bang Science Fair. As the demographic present was 9-18 year old students, teachers and parents, we were aware that we would have to have different activities and resources focused at the different ages.

June 22^nd

Log entry

For the second day of the Judd School trip the team led a synthetic biology talk for the students there and recorded a few more interviews. We received very positive feedback about the board game and the school asked for their own copy.

The morning lab team tested the cells that they cultured overnight, but the OD600 was lower than expected, and transformed the competent cells.

We also released part 1 of our ‘Meet the Team’ videos, signalling the start of our YouTube channel.

June 23^rd

Log entry

We checked on the competent cells transformed the previous day and all the colonies expressed strong fluorescence however the interlab parts had not worked.

During researching for data to put into our powerpoints for the Big Bang Science Fair we found that all the data on public perception of synthetic biology was rather out of date.

We decided to set up a synthetic biology survey which we intend to leave it running for the duration of our project and then analyse the data we have collected.

We Skyped Westminster and Purdue and now we’re setting up follow up talks with both teams to try and sort out potential collaborations.

June 24^th

Log entry

Most of the day was spent preparing presentations of our project ideas for an upcoming meeting with academics. We focused on the PAF idea, the genome integration idea and the kill switch idea. The viral-vector fungal killswitch idea was scrapped as there were too many potential flaws for us to iron out in the ten week process.

We met with last year’s Exeter iGEM team and discussed our ideas, what we’re doing for human practises and just general advice they wanted to give us.

June 27^th

Log entry

Professor Winlove and Dr Porter came to the academic meeting.They liked all three ideas and gave us feedback and advice. After discussing them the possible issues of each idea, we decided on a slightly expanded version of the kill switch idea as our project.

The rest of the day was split between researching more into kill switches and working on ideas for our stand at the Big Bang Science Fair, such as leaflets, posters, presentations, printed surveys and interactive activities to engage visitors.

June 28^th

Log entry

Today we have been preparing for the Big Bang Science Fair. We finished and printed off a poster showing how biology, chemistry, engineering and maths are all important in synthetic biology, a Synthetic Biology leaflet and an iGEM leaflet, the front and back of which are pictured below.

get imgs working

Also, we completed two presentations on Synthetic Biology and iGEM which we intend to run on a loop on two iPads for the duration of the fair.

The abstract for our project was written up and a basic form was uploaded to the Wiki. It focuses on how we can improve killswitches, as opposed to the initial killswitch idea of characterising the parts extensively. We intend to submit KillerOrange as our new part, and compare its functionality against that of KillerRed with a continuous culture. We also want to test if it is beneficial to have two killswitches in a system as opposed to one and look at the difference in stability of the killswitch if we integrate into into the chromosome.

June 29^th

Log entry

Today was the Big Bang Science Fair where the board game got a lot of interest and seemed very popular with the students who tried it out.

get imgs working

On top of that, we got the contact details of a home school teacher and a vice principle who are interested in potentially helping us set up focus groups for the board game. We also got a lot of new survey responses to add into our data collection on public perception on synthetic biology.

get img working

We started the interlab study today as our new parts arrived. We completed the transformation section of the study and left our prepared petri dishes in the incubator overnight.

June 30^th

Log entry

The agar plates we incubated overnight had colonies and they expressed GFP, as you can be seen in the photo below. The agar plates shown below are the competent cells containing plasmid 1, 2 and 3.

get img working

We released the video about the Big Bang Science Fair today, featuring interviews with several of the attendants there. We also started to write up the CRISPR proposal for the “CRISPR Design and DNA support” offered by Twist Bioscience and Desktop Genetics.

We skyped with Purdue for the second time and arranged a collaboration together. We have a Google Drive folder set up so it’s easier for us to communicate and request help from each other. Purdue will be running a continuous culture of a killswitch developed by Purdue’s previous iGEM team and sending us the data, and we’ll be helping them write up summaries of past team projects for their project database.

July 1^st

Log entry

Today was the third day of Interlab and our data so far is coming out as we expected.

We continued with the CRISPR proposal. We also skyped with UCL today, but due to how different our projects are, we had difficulty finding potential areas for collaboration.

July 4^th

Log entry

We discussed our ideas with Dr Paul James and told him that we had chosen to go with the kill switch idea and outlined the experiments that we wanted to do and began looking at parts for the constructs.

July 5^th

Log entry

Today we met with Dr Paul James and Dr Chloe Milner to recap our project idea and received feedback from them on the approach we should be taking towards it.

We met with Dr Nicky King again to discuss what we have so far for the synthetic biology module and gave us advice on different module structures we could look at and other academics we could talk to.

We had our headshots and team photo taken as the previous image sizes were not appropriate for the wiki.

get img working

July 6^th

Log entry

Today we attended the ‘Britain Needs Scientists’ fair. We we able to interview students and teachers about what they know about Synthetic Biology to further our human practices work. We had displayed our board game and edible DNA display which helped us to engage with more teachers about how to educate students on synbio.

After this we designed constructs for our KillerRed experiment choosing suitable parts from the iGEM registry.

July 7^th

Log entry

We recorded the very first episode of ‘Desert Island… Science?’ with Dr Nicky King today. We also had a meeting with Dr Sara Burton to discuss potential module structures and possible areas for funding.

We also skyped with UNL in the afternoon and looked at what experiments we are going to run.

July 8^th

Log entry

We have improved our board game, BioMech, for the upcoming school visit to Colyton Grammar School to trial the game.

We released the very first episode of ‘Desert Island… Science?’ today, with Dr Nicky King of the University of Exeter

July 11^th

Log entry

Today, we set up a collaboration with the Newcastle iGEM team. We hope to start running experiments to determine the thermal conductivity and convection effects in LB and M9 broth for them.

July 12^th

Log entry

We recorded and released the second episode of ‘Desert Island… Science?’ today. This time we interviewed Dr Sara Burton, a senior Biosciences lecturer at the University of Exeter. Dr Mark Ramsdale has also agreed to be interviewed. We have begun contacting people outside of the university so we can expand our series outside of Exeter.

We also had a Skype session with the UCC iGEM team today and agreed on a collaboration involving the testing of each of our parts.

July 13^th

Log entry

Today we continued with the Purdue collaboration, who wanted us to write summaries for their past project database. We also worked on designing parts and experiments.

July 14^th

Log entry

We met with Dr John Love who gave us advice on how we could expand our lab work more to looking at different ways to characterise the killswitches. We’ll be meeting with Dr Love and Dr James in sub teams next Tuesday to discuss our ideas in more depth.

July 15^th

Log entry

Today we received the 16 printed copies of our board game BioMech. We released a photo of the finalised board game on Twitter earlier, with this being the first time we’ve publically mentioned our board game.

We have also been in touch with The Sutton Trust in regards to the game, and they seem quite interested in the idea. Hopefully we will be able to sort something out with them in regards to widening participation.

July 18^th

Log entry

Today, four of our team members went on a school visit to Colyton Grammar School. They conducted a board game session with 26 students - our first board game session since having the final design printed. We made some surveys to give out to determine the impact of the board game on the students.

They also left behind three board games so the school could get extended use out of them, and continue to send us information on board game’ reception.

July 19^th

Log entry

We split into sub-teams to meet with Dr John Love and Dr Paul James separately and discuss progress in our respective areas.

July 20^th

Log entry

Today Professor Richard Kitney of Imperial College London agreed to partake in ‘Desert Island… Science?’.

We also received the board game survey data back from Colyton Grammar School and have done a basic analysis on the results - overall, 77.8% of students reacted positively, saying they had enjoyed the board game. We also got comments such as “It was a fun way for learning about synthetic biology.” and “It helped me remember most of the information because it was fun and engaging”.

July 21^st

Log entry

Today was taken up mostly by organising collaborations - we had a Skype session with Cardiff and talked about our projects and current progress. We also spoke with Westminster over Twitter about the possibility of collaborating.

We also started modelling today, researching the mechanism that Killer Red uses to lyse the cell.

July 22^nd

Log entry

The digestion-ligation transformation from yesterday did not work, so we will be repeating the digestion stage with RFP-tetracycline plasmids from Dr Chloe Singleton. For ligations, we made: promoter + RBS + destination, CDS + terminator + destination, and a control.

Yesterday’s PCR and gel electrophoresis were successful, but the products were not successfully transformed. We repeated the gel electrophoresis, which was successful, and transformed the products into E. coli DH5α cells which were left to incubate at 37°C overnight.

July 25^nd

Log entry

Professor Paul Freemont replied to our email request in regards to featuring on our interview series “Desert Island… Science?”. He said he would be keen to do so after his leave, so we will speak with him again in late August.

Transformations of ligations were successful. We made transformations of PCR products with a new KLD enzyme mix and plated all the transformations, and made glycerol stocks before doing minipreps.

July 26^th

Log entry

Today, we skyped with Dr Jon Marles-Wright for ‘Desert Island… Science?’, making this our first interview with an academic outside of the university. Dr Marles-Wright is one of the lead supervisors for the Edinburgh iGEM team.

We also decided to shift our project aim slightly. Instead of using Cas9, which we could not get to work, we will be looking genome integration instead, and comparing Killer Red and Killer Orange to ‘natural kill switches’ like hen egg Lysozyme and bovine DNAse1.

It was also a lab heavy day - we did digestion and ligation of promoter + RBS and CDS + terminator combinations in plasmids, and did transformations of ligation (1) (Promoter-RBS-CDS-Terminator in CAM), ligation (2) (Promoter-RBS-CDS-Terminator in CAM), positive control (RFP + chloramphenicol plasmid), negative control (WT competent cells) and destination (CAM destination plasmid).

July 27^th

Log entry

Andy and Alan worked in the Physics labs today, measuring the intensity of our light box, and calculating the W/cm^2 that the plates placed inside the light box will be receiving. We need to know this so we can look at how intensity impacts upon the rate at which Killer Red and Killer Orange kill cells.

Today’s lab team - Eloise, Emily, Hannah and Dan - started using a new method of cloning called Modular Cloning, which we intend to use codon optimised Killer Red and Killer Orange to submit to the registry. They also used an adjusted digestion and ligation method to cut RFP from template backbones, ran on a gel, then transformed for overnight incubation.

July 28^th

Log entry

The ligated Killer Red parts from yesterday were mini prepped, and the successful Cas9 colony was put in overnight incubation. We re-transformed the unsuccessful transformations from yesterday, as only the plasmids with chloramphenicol and tetracycline backbones were successful.

The successful Killer Red ligations were sent off for sequencing today to check that we made the part in the correct order sequence.

July 29^th

Log entry

Today marked a few additions to the modelling team due to the shift in project aims: Joel is continuing working on the KillerRed model, Andy will begin working on a hen egg Lysozyme model and Leanne will begin working on a bovine DNAse1 model. The current program of choice is SimBiology, an extension app on MATLAB, but that may change over the coming weeks depending on how compatible it is with our model aims.

We also released a new ‘Desert Island… Science?’ interview today, featuring Dr John Love. Dr Love is an associate professor of Synthetic Biology at the University of Exeter, and is our primary supervisor for iGEM.

The lab team for today - Jack, Emily, Joel and Hannah - did glycerol stocks, minipreps and qubits of the Cas9 part.

August 1^st

Log entry

Over the weekend, we released a short video on our interview with Richard Dawkins, who had come to Exeter as part of the International Society for Behavioral Ecology conference. The event, hosted by the university, brought academics from all across the globe to campus.

We had a new work experience student join us today called Jorge. He, like our previous work experience student Juliet, will be here with us for two weeks, helping out in the lab and generally operating as a standard team member.

Jorge was introduced to the full team during today’s team meeting, where we discussed the goals for this week inside and outside of the lab, and generally just checked up on the progress of each area of the project.

The lab team - Jack, Eloise, Pablo and Hannah - attempted to redo the unsuccessful plasmid backbone transformations from yesterday, but were not successful in amplifying the PCR product because the concentration was too low. However, some of the plasmid backbone transformations done today were successful, and have been overnighted so we can use them for cloning.

August 2^nd

Log entry

We swapped to using the QuickChange Multi kit instead of the Q5 kit for site-directed mutagenesis today as the Q5 kit had so far been unsuccessful, and the QuickChange kit could be used to mutate 5 sites at a time.

Today’s lab team also began using MoClo (Modular Cloning, mentioned in an earlier entry) to make codon optimised Killer Red and Killer Orange parts that we can submit to the registry. The products were transformed into S171 E. coli cells and overnighted. They also did some standard miniprepping and carried out qubits.

August 3^rd

Log entry

We put the order for our team t-shirts and hoodies/sweatshirts so they arrive in time for the Westminster meet-up. They have the iGEM logo on the front, our social media accounts on the back, and once we have a logo designed we’ll be sending them back to get the logo put on the front too!

Yesterday’s production of plasmid backbones was successful but the digestion and ligation of said backbones was not, so we repeated the digestion-ligation process today. The QuickChange multi kit also turned out to be unsuccessful, so we repeated that today to try remove multiple restriction sites in a pkD4 plasmid.

We transformed cloned Killer Red and Killer Orange in E. coli DH5α and overnighted them.

August 5^th

Log entry

We now have LB thermal conductivity data from the Newcastle collaboration experiments, and will be analysing them and sending them to the Newcastle iGEM team once we have completed the same for M9.

Joel began producing some fantastic logo ideas today, drawing from a moodboard produced by us a while back. Below is one of the team favourite logos - we plan to stick with the colour scheme featured, but may alter the design depending on further feedback.

Today in the lab, we made glycerol stocks of, miniprepped and carried out qubits of the overnights of Killer Red, Killer Orange and the plasmid backbones in DH5α and S171 strains that were made yesterday, and sent samples off for sequencing.

We also tried using the QuickChange multi kit again for site-directed mutagenesis, and the controls made indicated that we had been successful this time as they turned blue when tested.

August 10^th

Log entry

August 23^rd

@@ Line 3,066: / Line 3,066: @@
 			<div class="col-xs-12 div_log">
 				<h6 style="padding-left:0;padding-top:20px;">Log entry</h6>
+                                <p id="pp" style="padding-left:0;padding-right:0;padding-top:0;">
+<p id="pp">We swapped to using the QuickChange Multi kit instead of the Q5 kit for site-directed mutagenesis today as the Q5 kit had so far been unsuccessful, and the QuickChange kit could be used to mutate 5 sites at a time. </p>
+<p id="pp">Today’s lab team also began using MoClo (Modular Cloning, mentioned in an earlier entry) to make codon optimised Killer Red and Killer Orange parts that we can submit to the registry. The products were transformed into S171 E. coli cells and overnighted. They also did some standard miniprepping and carried out qubits.</p>
+                                </p>
 			</div>
 		</div>
@@ Line 3,094: / Line 3,097: @@
 				<h6 style="padding-left:0;padding-top:20px;">Log entry</h6>
 				<p id="pp" style="padding-left:0;padding-right:0;">
+                                      <p id="pp">We put the order for our team t-shirts and hoodies/sweatshirts so they arrive in time for the Westminster meet-up. They have the iGEM logo on the front, our social media accounts on the back, and once we have a logo designed we’ll be sending them back to get the logo put on the front too!</p>
 				</p>
 			</div>
 			<div class="col-xs-12 div_log">
 				<p id="pp" style="padding-left:0;padding-right:0;padding-top:0;">
+<p id="pp">Yesterday’s production of plasmid backbones was successful but the digestion and ligation of said backbones was not, so we repeated the digestion-ligation process today. The QuickChange multi kit also turned out to be unsuccessful, so we repeated that today to try remove multiple restriction sites in a pkD4 plasmid.</p>
+<p id="pp">We transformed cloned Killer Red and Killer Orange in E. coli DH5α and overnighted them.</p>
 				</p>
 			</div>
@@ Line 3,127: / Line 3,132: @@
 				<h6 style="padding-left:0;padding-top:20px;">Log entry</h6>
 				<p id="pp" style="padding-left:0;padding-right:0;">
+<p id="pp">We now have LB thermal conductivity data from the Newcastle collaboration experiments, and will be analysing them and sending them to the Newcastle iGEM team once we have completed the same for M9. </p>
 				</p>
 			</div>
 			<div class="col-xs-12 div_log">
 				<p id="pp" style="padding-left:0;padding-right:0;padding-top:0;">
+<p id="pp">Joel began producing some fantastic logo ideas today, drawing from a moodboard produced by us a while back. Below is one of the team favourite logos - we plan to stick with the colour scheme featured, but may alter the design depending on further feedback.</p>
+<p id="pp">Today in the lab, we made glycerol stocks of, miniprepped and carried out qubits of the overnights of Killer Red, Killer Orange and the plasmid backbones in DH5α and S171 strains that were made yesterday, and sent samples off for sequencing.</p>
+<p id="pp">We also tried using the QuickChange multi kit again for site-directed mutagenesis, and the controls made indicated that we had been successful this time as they turned blue when tested.</p>
 				</p>
 			</div>

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Revision as of 21:36, 19 October 2016

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