http://2012.igem.org/wiki/index.php?title=Special:Contributions&feed=atom&limit=50&target=RussellGritton2012.igem.org - User contributions [en]2024-03-28T13:36:07ZFrom 2012.igem.orgMediaWiki 1.16.0http://2012.igem.org/Team:NRP-UEA-Norwich/FrontTeam:NRP-UEA-Norwich/Front2012-11-24T01:19:52Z<p>RussellGritton: </p>
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<div>{{UEANRP}}<br />
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{{UEANRPLabs}}<br />
<br />
=Week 1=<br />
<br />
The first week of work was an exciting time for the NRP UEA iGEM team as we officially started our iGEM project! We spent the week largely focusing on planning out our project and taking part in some human practices activities before we can officially begin lab work. We were able to take part in the University of East Anglia School of BIO Colloquium, have talks on our project from PhD students working on similar topics, visit a fellow iGEM team and take part in a radio interview.<br />
<br />
==Day 1 (09/07/12)==<br />
<br />
Today the University of East Anglia School of Biological Sciences hosted its annual colloquium, an event where members of faculty gather together to discuss news over the year, give talks on their research and present posters. We were able to listen to many great talks on the research that has been going on in UEA and view the posters on display, where we were able to find out which researchers in the school were looking into nitric oxide and other topics relevant to our project, with a view to working with them in the future. We also spoke to representatives from Bioline, Qiagen, Fisher Science and Star Labs about the products they sold which we would likely require later on in the project; a few representatives even spoke about possible sponsorships and working with the iGEM team more closely, and we were able to leave with a few business cards giving us a few avenues to pursue.<br />
<br />
==Day 2 (10/07/12)==<br />
<br />
Today was a day largely involving meetings and planning ahead of the start of the project the following day. We met in the morning to discuss the project and we confirmed that we would be going into the lab the next day to transform ''E. coli'' with some earmarked BioBricks. Richard Bowater and Richard Kelwick told us that they had been able to synthesise a hybrid bacterial and mammalian, nitric oxide-sensitive promoter using PyeaR and NS2E9 elements in two orientations which would be the starting point of our BioBrick. We then looked at BioBricks produced by past teams to decide which BioBricks would be relevant to our project and therefore we would be transforming into the chassis; we decided on using the nitric oxide sensor developed by our advisors, and two BioBricks associated with bioluminescence.<br />
<br />
We also looked at deadlines for iGEM and began to draft our project outline for the deadline later that week. Richard Bowater also informed us of the COSHH forms we would be signing ahead of the lab work the next day and uploaded them to our Google Drive account so we could read over them and ensure our first day of lab work was completed with a high degree of safety. We then looked at the finances and began to discuss our trip to Amsterdam as well as organising the purchase of a Promega card for the university's Promega vending machine. The rest of the afternoon was then spent discussing our film, logo and future human practices events; we also found out that our radio interview would be in just 2 days!<br />
<br />
In the afternoon we sat in on a talk from a representative from the Biotechnology and Biological Sciences Research Council (BBSRC) on funding for research and how changes are being made to grants. We learnt that the BBSRC wanted their grant applicants to consider the 'impact' of their research (both in a scientific and economical sense) as part of their applications for funding; following this talk we too will be looking at what impact our research may make as part of our work on the ethics and future applications of our work.<br />
<br />
==Day 3 (11/07/12)==<br />
<br />
===Meetings===<br />
<br />
[[File:NRPUEARabbit.jpg | right | thumb | Our original rabbit-based logo]]<br />
<br />
Today was our first day of lab work, however, before we could do that there were more meetings to be had and decisions to be made! We discussed the concept logos that our artist, Amy Congdon, had produced. Many of them contained the image of a rabbit, an image synonymous with UEA's campus, and an image found on this very wiki. We felt perhaps that because our logo would be very much our public image, and that outside of the iGEM world a logo baring a rabbit prominently may give the wrong impression of synthethic biology to the public, that the rabbit aspect of the logo may be better as a smaller piece of the image, or even non-existent in it. We did, however, believe that the "N=ORWICH" part of the logo was extremely important and clever in representing our project on nitric oxide , and this can be seen in the finalised version of the logo all over the wiki!<br />
<br />
We also focused on the information we wanted to present in our interview on STAR Radio the next day. As we were told we would be on both the farming and the business portions of the show, we focused on how to discuss Synthetic Biology in the senses of both agriculture and business. Further to this, we ensured that we had a clear and precise definition of synthetic biology in order to help put the idea across to the radio show's listeners and hopefully publicise the branch of science and help dispel fears about it. <br />
<br />
In the afternoon we had our end of day lab meeting where we built on the project outline for Sunday's iGEM deadline and contacted Cambridge's iGEM team to discuss a visit and meeting while we were in Cambridge for the radio interview.<br />
<br />
===Labs===<br />
<br />
Today was our first day in the labs and we began by transforming NEB 5-alpha ''Escherichia coli'' with three BioBricks we had picked out as being applicable to our project in order to characterise them and investigate how far we would be able to use them in our project. When choosing the BioBricks we looked at those that had been used in previous years to either sense/report on nitric oxide specifically, or those that been used in sensing/reporting on physiologically relevant molecules that we could adapt for nitric oxide sensation. The three BioBricks we used were:<br />
<br />
<br />
. [http://partsregistry.org/Part:BBa_K381001 BBa_K381001]; produced by Bristol in 2010, a nitrite and nitrate sensitive promoter (PyeaR) using GFP to report nitric oxide presence. PyeaR is usually repressed by NsrR, however when nitrates are converted to nitric oxide, NsrR is sequestered thus preventing its repression of PyeaR and allowing the production of GFP. We can therefore, utilise this function in order to show levels of nitric oxide via GFP expression.<br />
<br />
<br />
. [http://partsregistry.org/Part:BBa_K325909 BBa_K325909]; produced by Cambridge in 2010, a reporting BioBrick using L-arabinose to induce the pBADpromoter, which in turn activates the lux operon resulting in bioluminescence. We believe characterisation of this BioBrick may allow us to build a circuit along with nitric oxide-sensing BioBricks to result in a standard part that becomes bioluminescent in the presence of nitric oxide.<br />
<br />
<br />
. [http://partsregistry.org/Part:BBa_K325100 BBa_K325100]; produced by Cambridge in 2010, a reporting BioBrick using L-arabonose to induce the pBAD promoter, which in turn activates luciferase and luciferin regenerating enzyme, resulting in bioluminescence. We believe characterisation of this BioBrick may allow us to build a circuit along with nitric oxide-sensing BioBricks to result in a standard part that becomes bioluminescent in the presence of nitric oxide.<br />
<br />
<br />
Protocol on our first transformation can be found [[Team:NRP-UEA-Norwich/Protocol | Here]]<br />
<br />
<br />
We made an amendment to the original protocol that saw double the amount of cells being used in the experiment in order to maximise our number of transformed cells at the end. The agar plates with transformed bacteria on them were incubated over night.<br />
<br />
==Day 4 (12/07/12)==<br />
<br />
===Cambridge===<br />
<br />
[[File:NRPUEACambridge.JPG | right | thumb | The NRPUEA Team meeting the Cambridge team]] <br />
<br />
Today was the day of our road trip to Cambridge where we would be meeting the Cambridge iGEM team and visiting their labs as well as attending 'The Business Hub' program om STAR radio to give an interview on iGEM and synthetic biology. Before leaving the UEA Campus we finalised everything for the interview and gave mock interviews to Khadija and Pascoe, the two members of the team that would be speaking on air, in order to help prepare for the kinds of questions they may be asked. We also organised what our aims were for our meeting with Cambridge's iGEM team, and decided that as their project appeared to be of a similar like to ours that we would look into collaborating with one another.<br />
<br />
We arrived at Cambridge's Downing Site, where their iGEM team were working in the Plant Sciences building, and were taken straight up to the lab to meet the team. In the lab we were introduced to their members and discussed our projects and the various problems we had encountered throughout the experimental side of the competition. Their team were a couple of weeks ahead of ours so it was interesting to hear their advice on certain aspects of the lab work; in addition we exchanged thoughts and constructive criticism on regards to each others projects. Ultimately we felt that both teams benefitted from the meeting and gain valuable information and experience.<br />
<br />
Later in the day we traveled to STAR Radio 107.9/1 FM for our interview on their 'The Business Hub' show. The presenter met us and was extremely interested in the competition as well as synthetic biology as a whole. Khadija and Pascoe conducted the interview and did a fantastic job; the interview went very well and we were able to put across a lot of information on synthetic biology, iGEM and our own project. We hope that once the programme airs that we will receive feedback from members of the public.<br />
<br />
===Labs===<br />
<br />
After checking the plates of transformed ''E. coli'' we found that none had grown. We theorised that the chloramphenicol concentration of the agar might have been to high (50µg/ml) and decided to spend the rest of the week researching the growing conditions for the bacteria, the details of the cells we were using as well as compare various protocols of transformation used and developed by other iGEM teams in order to perform a successful transformation of ''E. coli'' again the next Monday.<br />
<br />
==Day 5 (13/07/12)==<br />
<br />
===Human Practices===<br />
<br />
Our team artist, Amy Congdon, visited today in order to discuss our video to present at the Jamboree in Amsterdam as well as to discuss the design aspects of our project. The next Friday we will be going into a school in order to present about synthetic biology and the project, therefore Amy helped out the members of the team involved in that presentation to produce some equipment and props to aid with it. She also looked over our wiki designs and adapted the logo with our suggestions in order to produce a final draft of the logo that we all felt was the most suitable one. Amy's ideas on the video were extremely exciting and captured the entire team's imaginations as to our project and how we will be presenting it and we all began to anticipate starting to work on it. She also gave us some fantastic advice on the safety and ethics aspect of the project, as well as suggesting creative ways of addressing the issues involved with each.<br />
<br />
===Research===<br />
<br />
Those that did not meet with Amy instead focused on researching what had gone wrong with growing the ''E. coli'' and how it could be fixed, as well as looking at other BioBricks we could use and how to build the project on further once we had characterised some of the relevant BioBricks and produced our own. Having looked at previous protocols we felt that our agar plates contained too much chloramphenicol and therefore we decided it would be best to reduce the amount when trying to transform and grow the bacteria during the next week. We also looked at how previous teams had worked on creating data from BioBricks that sensed and reported on the presence of physiological molecules, and researched into how we could use expressions such as fluorescence or bioluminescence to determine the levels of nitric oxide present in an environment.<br />
<br />
In the afternoon we were given a presentation by Sebastian Runkel, a PhD student at UEA whose research specialises in nitric oxide. He helped us to understand how nitric oxide was used in the body and what the major limitations and obstacles would be in working with it, obstacles and limitations we then went on to discuss and investigate how we could overcome them. Sebastian also spoke about using Salmonella as a potential chassis instead of ''E. coli'' due to its higher competency with nitrogen species, and we went on to discuss this as a possible path to investigate once we were able to produce a BioBrick in the ''E. coli'' chassis. This lead to new ideas on how our project might be utilised in the future.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-NorwichTeam:NRP-UEA-Norwich2012-09-27T00:57:37Z<p>RussellGritton: </p>
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<a href="http://www.norwichresearchpark.com/home.aspx"><img src="https://static.igem.org/mediawiki/2012/d/dc/NRPF1.png"></a><img src="https://static.igem.org/mediawiki/2012/0/0c/NRPF2.png"><a href="https://igem.org/Main_Page"><img src="https://static.igem.org/mediawiki/2012/7/7e/NRPF3.png"></a><img src="https://static.igem.org/mediawiki/2012/d/d8/NRPF4.png"><a href="http://www.uea.ac.uk/"><img src="https://static.igem.org/mediawiki/2012/a/a1/NRPF5.png"></a><br />
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</html></div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-NorwichTeam:NRP-UEA-Norwich2012-09-27T00:57:11Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRPHome}}<br />
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<a href="http://www.norwichresearchpark.com/home.aspx"><img src="https://static.igem.org/mediawiki/2012/d/dc/NRPF1.png"></a><img src="https://static.igem.org/mediawiki/2012/0/0c/NRPF2.png"><a href="https://igem.org/Main_Page"><img src="https://static.igem.org/mediawiki/2012/7/7e/NRPF3.png"></a><img src="https://static.igem.org/mediawiki/2012/d/d8/NRPF4.png"><a href="http://www.uea.ac.uk/"><img src="https://static.igem.org/mediawiki/2012/a/a1/NRPF5.png"></a><br />
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</html></div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/ProjectTeam:NRP-UEA-Norwich/Project2012-09-27T00:55:59Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRP}}<br />
<br />
{{UEANRPProjects}}<br />
<br />
==NRP-UEA-Norwich Project Overview==<br />
<br />
Sensory BioBrick systems have been a large constituent of previous iGEM projects in which teams have combined impressive amounts of logic with limitless creativity in order to produce synthetically engineered organisms with the ability to detect the presence of specific substrates; this was achieved by combining various promoters and reporters to produce novel gene systems of great breadth and depth.<br />
<br />
We too have taken a sensory approach to our project and have produced systems involved in the sensation of nitric oxide (NO). Originally we set out to develop a bacterial and mammalian hybrid NO-sensing promoter (which we have achieved); we then looked into ways of quantifying the levels of highly reactive and difficult to measure NO within a system, leading to us producing a novel gene regulation system known as the comparator circuit. Throughout the project we went on to look at theoretical alternative approaches to the gene systems we have produced.<br />
<br />
====Overall in our project we have produced 6 sensory BioBricks, 2 BioBricks involved in gene regulation, and have further characterised 4 more BioBricks. All 8 of our BioBricks have been submitted to the registry and the 6 sensory BioBricks have been characterised. Our project consists of three components, read their summaries on this page or click on the images below to see more detailed information including our BioBrick characterisation data.====<br />
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{| style="color:#1b2c8a;background-#6C0;" cellpadding="3" cellspacing="1"width="62%" align="center"<br />
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!align="left"|[[File:NRPNOLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing |300px]]<br />
!align="left"|[[File:NRPCompLogo.png |link=https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit |300px]]<br />
!align="left"|[[File:NRPTheoreticalLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/TheoreticalProjects |300px]]<br />
|-<br />
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!align="left"|6 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 BBa_K774000] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 BBa_K774001] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 BBa_K774005] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 BBa_K774006] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004]<br />
!align="left"|2 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774002 BBa_K774002] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774003 BBa_K774003]<br />
!align="left"|Theoretical BioBricks: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774100 BBa_K774100]<br />
|-<br />
|<br />
|}<br />
<br />
====Existing iGEM BioBrick parts which we characterised during our project:====<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 PyeaR Promoter BBa_K216005] -- [http://partsregistry.org/Part:BBa_K381001 PyeaR Promoter + GFP BBa_K381001] -- [http://partsregistry.org/Part:BBa_E0420 enhanced Cyan Fluorescent Protein + RBS + Terminators BBa_E0420] -- [http://partsregistry.org/Part:BBa_K081014 Red Fluorescent Protein + RBS + Terminators BBa_K081014]<br />
<br />
----<br />
<br />
[[File:NRPNOLogo.png | centre | link=Team:NRP-UEA-Norwich/NOSensing]]<br />
<br />
<br />
Our main project has resulted in the production of a hybrid bacterial and mammalian promoter optimised for induction by nitric oxide, nitrates and nitrites. We have ligated PyeaR, a known bacterial promoter [http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005] (Cambridge 2009) in the parts registry, with its mammalian counterpart, CArG. The resulting hybrid promoter has been synthesised in two orientations; PyeaR (bacterial, B) upstream of CArG (mammalian, M), nicknamed (B-M); and CArG upstream of PyeaR (M-B). These orientations were submitted to the parts registry as our first two BioBricks.<br />
<br />
Each orientation of the promoter was ligated to enhanced cyan fluorescent protein (eCFP) and red fluorescent protein (RFP) to produce four new BioBricks which have been submitted to the parts registry. These promoter + fluorescent protein BioBricks have been characterised following transformation into ''Escherichia coli'' and induction by potassium nitrate using methods such as flow cytometry, fluorescence-activated cell sorting (FACS) and scanning with a fluorometer. The data from these experiments has proved that our promoter works as we expected it to. <br />
<br />
We have also transfected M-B + eCFP into a human breast cancer cell line, MCF7, and have provided data to show that the system is flexible and can be used in both eukaryotes and prokaryotes. We believe the promoters we have produced have relevant uses in cancer therapeutics, soil fertilisation and detection of emissions from industries such as construction.<br />
<br />
Our 6 BioBricks:<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian/B-M (PyeaR-CArG) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial/M-B (CArG-PyeaR) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP]<br />
<br />
----<br />
<br />
<br />
[[File:NRPCompLogo.png | centre | link=Team:NRP-UEA-Norwich/ComparatorCircuit]]<br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
The lack of specificity of the bacterial promoter, pYeaR, used in the hybrid promoter was a pitfall that was always a concern when it came to accurately sensing nitric oxide. From this potential problem spawned a potential solution; the Comparator Circuit. This pair of BioBricks are designed to specifically bind to each other while ligated to two different promoters of overlapping specificity to result in an integration of the conflicting outputs of the two opposing gene systems.<br />
<br />
<br />
Our system relies on two constructs that interact via complimentary base pair sequences both before and after the ribosome binding site of the reporter protein. The idea being that, when both transcripts are present in the chassis, they would bind together, inhibiting the translation of the reporter proteins.<br />
<br />
<br />
Any imbalance of transcription due to the presence of the substrate of interest results in free mRNA of the gene system that detects that substrate. Crucially, if both promoters detect the same substrates but differ with one extra substrate being detected by one of the promoters, it is this substrate and this substrate only that our system will be able to detect in a simple and quantitative way. <br />
<br />
<br />
Our team have constructed a countercurrent comparator circuit in which the reporter proteins are at the same end of the complimentary region, although a contracurrent system has been theorised. Both systems share a crucial subtractive nature comparable to an analogue computer. We envisage that, should the system be fine-tuned and expanded on, a variety of different business sectors from agriculture to spinoff pharmaceutical companies (<html><font size=2pt><b>Please check out our <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details</b></font></html>) could capitalise on this novel genetic technology.<br />
<br />
<br />
What we have produced is a BioBrick pair that work in harmony, when ligated to promoters of interest and genes of interest, to sequester translation when both mRNA transcripts are present in the cell. The use of quantative tuners with these BioBricks is encouraged to ensure that the transcription rate both gene constructs are equal when both promoters are transcribing at their optimal rate. Although the parts have been submitted to the registry and theoretically characterised, time constraints have meant that further lab-based characterisation could not occur.<br />
<br />
<br />
However, we hope to utilise any free time in our timetables sduring the next semester to characterise the BioBricks further (please see our project proposal), and hope that we will be given a chance to present our further findings at MIT! <br />
<br />
<br />
To conclude, what we have created is a pair of antagonistic BioBricks that turned the pair of mRNAs in which they reside into translational repressor molecules when both are transcribed in tandum within a specific chassis of interest, a new application for mRNA complimentary base pairing within the registry and a project we feel could go very far indeed.<br />
<br />
<br />
----<br />
<br />
<br />
[[File:NRPTheoreticalLogo.png | centre | link=Team:NRP-UEA-Norwich/TheoreticalProjects]]<br />
<br />
<br />
In addition to the two main projects, we have also worked to elaborate on some of the teams earlier ideas during the project:<br />
<br />
'''Multiplicative circuit'''<br />
<br />
The comparator circuit that we have created integrates two different transcription levels in a negative manner (subtraction) to perform a range of functions a range of different integrations (calculations) are necessary to this end we have also designed a system that would allow one signal (transcription rate) to be divided by the transcription level of another promoter. To achieve this we have used the processes of attenuation and the three loop system (in the trp leader). the system has also been mathematically modeled. To find out more please click the image above. <br />
<br />
<br />
'''Multisensor'''<br />
<br />
There are many groups of chemical species for which there are no current biological techniques for distinguishing between each of these species and quantitatively analysing its concentration. Here we outline a possible approach for solving this problem using non-specific transcription factors and promoters. We use nitrates, nitrites and nitric oxide as our example group. To find more please click the image above.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/ProjectTeam:NRP-UEA-Norwich/Project2012-09-27T00:55:25Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRP}}<br />
<br />
{{UEANRPProjects}}<br />
<br />
==NRP-UEA-Norwich Project Overview==<br />
<br />
Sensory BioBrick systems have been a large constituent of previous iGEM projects in which teams have combined impressive amounts of logic with limitless creativity in order to produce synthetically engineered organisms with the ability to detect the presence of specific substrates; this was achieved by combining various promoters and reporters to produce novel gene systems of great breadth and depth.<br />
<br />
We too have taken a sensory approach to our project and have produced systems involved in the sensation of nitric oxide (NO). Originally we set out to develop a bacterial and mammalian hybrid NO-sensing promoter (which we have achieved); we then looked into ways of quantifying the levels of highly reactive and difficult to measure NO within a system, leading to us producing a novel gene regulation system known as the comparator circuit. Throughout the project we went on to look at theoretical alternative approaches to the gene systems we have produced.<br />
<br />
====Overall in our project we have produced 6 sensory BioBricks, 2 BioBricks involved in gene regulation, and have further characterised 4 more BioBricks. All 8 of our BioBricks have been submitted to the registry and the 6 sensory BioBricks have been characterised. Our project consists of three components, read their summaries on this page or click on the images below to see more detailed information including our BioBrick characterisation data.====<br />
<br />
<br />
<br />
{| style="color:#1b2c8a;background-#6C0;" cellpadding="3" cellspacing="1"width="62%" align="center"<br />
|-<br />
|<br />
!align="left"|[[File:NRPNOLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing |300px]]<br />
!align="left"|[[File:NRPCompLogo.png |link=https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit |300px]]<br />
!align="left"|[[File:NRPTheoreticalLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/TheoreticalProjects |300px]]<br />
|-<br />
|<br />
!align="left"|6 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 BBa_K774000] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 BBa_K774001] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 BBa_K774005] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 BBa_K774006] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004]<br />
!align="left"|2 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774002 BBa_K774002] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774003 BBa_K774003]<br />
!align="left"|Theoretical BioBricks: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774100 BBa_K774100]<br />
|-<br />
|<br />
|}<br />
<br />
====Existing iGEM BioBrick parts which we characterised during our project:====<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 PyeaR Promoter BBa_K216005] -- [http://partsregistry.org/Part:BBa_K381001 PyeaR Promoter + GFP BBa_K381001] -- [http://partsregistry.org/Part:BBa_E0420 enhanced Cyan Fluorescent Protein + RBS + Terminators BBa_E0420] -- [http://partsregistry.org/Part:BBa_K081014 Red Fluorescent Protein + RBS + Terminators BBa_K081014]<br />
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[[File:NRPNOLogo.png | centre | link=Team:NRP-UEA-Norwich/NOSensing]]<br />
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Our main project has resulted in the production of a hybrid bacterial and mammalian promoter optimised for induction by nitric oxide, nitrates and nitrites. We have ligated PyeaR, a known bacterial promoter [http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005] (Cambridge 2009) in the parts registry, with its mammalian counterpart, CArG. The resulting hybrid promoter has been synthesised in two orientations; PyeaR (bacterial, B) upstream of CArG (mammalian, M), nicknamed (B-M); and CArG upstream of PyeaR (M-B). These orientations were submitted to the parts registry as our first two BioBricks.<br />
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Each orientation of the promoter was ligated to enhanced cyan fluorescent protein (eCFP) and red fluorescent protein (RFP) to produce four new BioBricks which have been submitted to the parts registry. These promoter + fluorescent protein BioBricks have been characterised following transformation into ''Escherichia coli'' and induction by potassium nitrate using methods such as flow cytometry, fluorescence-activated cell sorting (FACS) and scanning with a fluorometer. The data from these experiments has proved that our promoter works as we expected it to. <br />
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We have also transfected M-B + eCFP into a human breast cancer cell line, MCF7, and have provided data to show that the system is flexible and can be used in both eukaryotes and prokaryotes. We believe the promoters we have produced have relevant uses in cancer therapeutics, soil fertilisation and detection of emissions from industries such as construction.<br />
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Our 6 BioBricks:<br />
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[http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian/B-M (PyeaR-CArG) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial/M-B (CArG-PyeaR) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP]<br />
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[[File:NRPCompLogo.png | centre | link=Team:NRP-UEA-Norwich/ComparatorCircuit]]<br />
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[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
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The lack of specificity of the bacterial promoter, pYeaR, used in the hybrid promoter was a pitfall that was always a concern when it came to accurately sensing nitric oxide. From this potential problem spawned a potential solution; the Comparator Circuit. This pair of BioBricks are designed to specifically bind to each other while ligated to two different promoters of overlapping specificity to result in an integration of the conflicting outputs of the two opposing gene systems.<br />
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Our system relies on two constructs that interact via complimentary base pair sequences both before and after the ribosome binding site of the reporter protein. The idea being that, when both transcripts are present in the chassis, they would bind together, inhibiting the translation of the reporter proteins.<br />
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Any imbalance of transcription due to the presence of the substrate of interest results in free mRNA of the gene system that detects that substrate. Crucially, if both promoters detect the same substrates but differ with one extra substrate being detected by one of the promoters, it is this substrate and this substrate only that our system will be able to detect in a simple and quantitative way. <br />
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Our team have constructed a countercurrent comparator circuit in which the reporter proteins are at the same end of the complimentary region, although a contracurrent system has been theorised. Both systems share a crucial subtractive nature comparable to an analogue computer. We envisage that, should the system be fine-tuned and expanded on, a variety of different business sectors from agriculture to spinoff pharmaceutical companies (<html><font size=2pt><b>Please check out our <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details</b></font></html>) could capitalise on this novel genetic technology.<br />
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What we have produced is a BioBrick pair that work in harmony, when ligated to promoters of interest and genes of interest, to sequester translation when both mRNA transcripts are present in the cell. The use of quantative tuners with these BioBricks is encouraged to ensure that the transcription rate both gene constructs are equal when both promoters are transcribing at their optimal rate. Although the parts have been submitted to the registry and theoretically characterised, time constraints have meant that further lab-based characterisation could not occur.<br />
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However, we hope to utilise any free time in our timetables sduring the next semester to characterise the BioBricks further (please see our project proposal), and hope that we will be given a chance to present our further findings at MIT! <br />
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To conclude, what we have created is a pair of antagonistic BioBricks that turned the pair of mRNAs in which they reside into translational repressor molecules when both are transcribed in tandum within a specific chassis of interest, a new application for mRNA complimentary base pairing within the registry and a project we feel could go very far indeed.<br />
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[[File:NRPTheoreticalLogo.png | centre | link=Team:NRP-UEA-Norwich/TheoreticalProjects]]<br />
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'''Multiplicative circuit'''<br />
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The comparator circuit that we have created integrates two different transcription levels in a negative manner (subtraction) to perform a range of functions a range of different integrations (calculations) are necessary to this end we have also designed a system that would allow one signal (transcription rate) to be divided by the transcription level of another promoter. To achieve this we have used the processes of attenuation and the three loop system (in the trp leader). the system has also been mathematically modeled. To find out more please click the image above. <br />
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'''Multisensor'''<br />
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There are many groups of chemical species for which there are no current biological techniques for distinguishing between each of these species and quantitatively analysing its concentration. Here we outline a possible approach for solving this problem using non-specific transcription factors and promoters. We use nitrates, nitrites and nitric oxide as our example group. To find more please click the image above.<br />
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In addition to the two main projects, we have also worked to elaborate on some of the teams earlier ideas during the project.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/NotebookTeam:NRP-UEA-Norwich/Notebook2012-09-27T00:46:20Z<p>RussellGritton: </p>
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<div>{{UEANRP}}<br />
<br />
{{UEANRPLabs}}<br />
<br />
The NRP-UEA team was funded by the Wellcome trust for 10 weeks commencing from July 9th to September 14th. Leading up to the start day, the team met up to discuss the scope, the runnings and the time frame within which we had to complete a synthetic biology project. It was during that month leading up to the start date that we decided to build a sensor for nitric oxide and the rest was history, or at least '''slowly''' fell into place.<br />
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<br />
At the start of the 10 weeks, we were a little lost as what to do. Everyone had really big ideas as how to carry out the project but relatively little experience in research. Having been a little molly coddled in lab practice, we were suddenly doing everything ourselves, finding solutions and using research to solve problems. Needless to say, it took us many weeks to get into the swing of things. As emphasised, things kick started rather slowly. <br />
<br />
<br />
Initial problems included transforming iGEM BioBricks, DNA isolation, gel purification and ligations. In general we encountered many challenges in all steps of cloning. However, with jokes and encouragement from each other, we worked on the finicky details of the protocols, such as a flick here, a centrifuge short spin there and lower antibiotic resistance there. Slowly but steadily we improved our lab skills. Around week 4 to 5, lab work began to fall piece by piece into place. It was also around this time that our grand ideas and designs for building a quantitative and specific modular sensor came together. All in all by the half way point, things were looking up. We had successfully cloned BM and MB into the pSB1C3 iGEM backbone and isolated the DNA. We had prepared reporter proteins: RFP (BBa_R0080) and eCFP (BBa_E0420) for ligation with the potential BioBricks. The constructs were also ready to be sent off for synthesis.<br />
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Onto the latter half of the project, we still had much to do. With our potential first BioBricks, we set about planning to ligate reporter proteins to them to both improve the BioBricks and characterise them. We also set started to further characterising other reporter such as PyeaR + GFP.<br />
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RFP and CFP BioBricks that we had transformed, were selected and prepared to be ligated to BM and MB. Possibly due to denaturing, contamination and mutation, the eCFP and RFP DNA we had hydrated was not transforming and so we decided to use other BioBricks. However none of the ones on our plates had ribosome binding sites and confirmed sequences. Therefore, we streaked the DNA we had from the original transformation plates of RFP and CFP and restarted the cloning process. Towards the end of the allotted 10 weeks, we finally cloned the reporter proteins into our BioBricks and characterised them. <br />
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<br />
The growth study characterisations we carried out involved measuring the affects of nitrates on PyeaR + GFP transformed cells and the affect PyeaR + GFP, BM and MB Biobricks on the growth rate of ''E. coli''. The results of these were at times what we expected and at times not. <br />
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In week 9, the synthesised DNA of our constructs arrived! Despite this being later than we originally had hoped for, we were determined to make things work and immediately set off to get started on the cloning process. By this point, everyone was very rehearsed in the methods of cloning and almost everything worked first time. It was also through the cloning of our constructs that new champions of miniprepping arose. By the end of week 10, we had successfully cloned Construct 1 and 2 into pSB1C3 and sent the DNA off for sequencing and to iGEM. <br />
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To finish the characterisation process off, we continued lab work into week 11, where we were able to use FACs, flow cytometry and a fluorometer to gain insight of the functionality of the new Biobricks. Besides these tools, we also transfected BM and MB into mammalian cells, prooving that the combination of mammalian and bacterial promoters brought about flexibility of chassis. Using these, we were able to fully characterise BM and MB.<br />
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<br />
For full details of the lab work carried out, check out the individual weeks. The details of the protocols used are stated and also the full details of experiments and results can be found in the experiments page.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/Human_OutreachTeam:NRP-UEA-Norwich/Human Outreach2012-09-27T00:36:34Z<p>RussellGritton: /* The Future of Science event held at the forum, Norwich (Sunday 19th August) */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Human Practices=<br />
<br />
<br />
The UEA NRP team really went up and beyond this year, carrying out a wide selection of activity's to inform the public of both synthetic biology and iGEM, aiming to raise awareness of this branch of science. We used many methods of communication in order to cater for a range of audiences, including a radio show, newspaper articles, pubic events and talks as well as collaborating with all the other UK teams, in order to organize the UK team meet up and steam it live on the internet. <br />
<br />
== iGEM collaborations==<br />
<br />
===The iGEM UK team meet up, hosted by the NRP-UEA team at Google campus London (Friday 17th August) The most open and publically accessible UK team meetup EVER===<br />
<br />
[[File:Hangout stats.png | center]]<br />
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[[File:UKiGEMteams.jpg | 300px | right]]<br />
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The NRPUEA iGEM team organised and hosted the UK team meet up at Google campus in London. The NRPUEA iGEM team greeted the UK teams, provided refreshments and a lovely buffet lunch, as well as chairing the speeches and making sure the day ran smoothly. There was a great atmosphere at the meet up, and the event was a huge success.<br />
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The day consisted of a number of guest speakers, including advice and tips for iGEM success from Dr. Tom Ellis (advisor of the 2009 winning iGEM project E.chromi) and an interactive talk on synthetic biology and science communication from the BBC science presenter, Nature editor and Guardian writer, Dr. Adam Rutherford. The UK iGEM teams presented a poster and a 15 minute presentation about their project. This gave the members of each team the chance to practice their presentation skills before the European team meet up, as well as a chance for others to ask questions and see what other project routes the other teams have taken. <br />
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The event was also available '''LIVE on the internet''' via Google+ hangout, to allow the public and those that were unable to attend to still watch the presentations. Despite there being a few technical difficulties with the quality of the image, figures have shown that over 600 people watched the event.<br />
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[https://dl.dropbox.com/u/9957127/UK%20iGEM%20Team%20Hangout%2017th%20Aug%202012%20Programme.pdf '''Click here for programme PDF''']<br />
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<br />
=== Communicating with other iGEM teams===<br />
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As part of our efforts to collaborate with the other iGEM teams this year we've made an effort to keep in contact with an array of iGEM teams. Here are some highlights:<br />
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- We communicated with all of the UK iGEM teams via Twitter, Email, Facebook and Google+ before, during and after the UK team meet up.<br />
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- We tweeted with many other iGEM teams from around the world and closely followed the iGEM community.<br />
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- We video called the St Andrews iGEM team via Google+ Hang-out to discuss our projects further as well as to seek advice about the arrangements for the UK team meet-up.<br />
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- Our advisor Richard Kelwick was interviewed by the Edinburgh iGEM team via Skype about our teams perspectives on public awareness and understanding of synthetic biology.<br />
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- We visited the Cambridge team near the beginning of our project and they were kind enough to give us a tour of their labs. <br />
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- Finally, the British Columbia team sent us a questionnaire about intellectual property that we happily filled out.<br />
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==Public engagement==<br />
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===The Future of Science event held at The Forum, Norwich (Sunday 19th August)===<br />
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[[File:Forum_team.JPG | 300px| right]]<br />
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The Forum is located at the heart of the city of Norwich, and hosts a wide range of free events and exhibits to be enjoyed by the whole community. The venue is packed with restaurants, cafes and shops, as well as being home to Norwich library and BBC East Anglia. <br />
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[[File:Poster for Forum.jpg | 250px | left]]<br />
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The NRPUEA iGEM team were fortunate enough to hold an interactive day at the Forum, giving the public a chance to engage within the world of synthetic biology. Before the date of the event, the team increased awareness via a range of techniques, including posters, leaflets and newspaper and internet adverts, including the Forums website. This resulted with a large turnout at the event, including the local press. <br />
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At the Future of Science event, the team presented a selection of informative posters, revealed their new film on a large screen, as well as showing examples of work carried out by the students during iGEM. For example, the public could view ''E.coli'' which had been transformed with GFP within a UV light box, as well as carrying out a counting colonies exercise. The NRPUEA team also created a range of synthetic biology educational material for the event, to inform the public of the concepts and terminology of synthetic biology. The different activities were designed to interest a selection of different ages. The educational resources include a range of quizzes and worksheets such as crosswords and matching exercises.<br />
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[[File:Child_making_plasmid.jpg |300px | right]]<br />
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For younger children we designed a colourful jigsaw plasmid (as seen in image). The idea of this activity was for the participants to choose jigsaw pieces that represented the genes that code for certain characteristics, such as green or red hair, and piece them together to form a complete plasmid. Once they had designed their plasmid they used a biscuit base, icing of different colours and various sweets to create a "biscuit creature" that corresponded to the plasmid they had created. The learning outcome of this activity was to give the children an understanding that a different selection of genes, results in different characteristics, and that editing the DNA can result in a change in the characteristics. The activity was very popular and most of the participants seemed to understand this concept, as well as having great fun.<br />
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The event attracted a range of ages, as well as mixed levels of previous interest into synthetic biology. It also became clear that many members of the public knew very little about synthetic biology, and therefore were very interested in finding out more about the new branch of biology and the iGEM competition. The NRPUEA iGEM team also took the opportunity to talk to the public and listen to their opinions about the work the iGEM teams are carrying out, which demonstrated how mixed the view of the public were. The day was a huge success.<br />
<br />
'''Synthetic Biology activity sheets'''<br />
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[https://static.igem.org/mediawiki/2012/1/1f/Word_Search_of_Synthetic_Biology.pdf Synthetic Biology Wordsearch],<br />
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[https://static.igem.org/mediawiki/2012/3/3e/Decode_Disney.pdf Disney DNA Decoder],<br />
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[https://static.igem.org/mediawiki/2012/e/e5/Questions_comprehensive.pdf Match The Word To Its Definition],<br />
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[https://static.igem.org/mediawiki/2012/8/8e/Decode_Music_Artists_and_Films.pdf Music Artists & Films DNA Decoder]<br />
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=== Creation of a futuristic company, Quanticare and production of a thought-provoking film===<br />
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[[File:Tattoo1.png |300px]][[File:QuanticareLogo_(2).png |300px]][[File:Tattoo2.png |300px]]<br />
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As part of our human practices we decided to look into an artistic approach to bringing synthetic biology to a wider audience. As part of this we created "Quanticare", a fictitious and futuristic company forming on the back of a rise to prominence of synthetic biology over the next few years, as well as the results of our own projects on nitric oxide sensing and the comparator circuit. The fictitious company's latest development is to introduce a visual bio sensors in to the human body in the form of a tattoo-based health monitor, ''Cura''. <br />
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Not only did the development of the fictitious company allow the team to explore potential future applications of synthetic biology and bio sensors, but also via the creation of a film , fuel a range of ethical questions. The team released the video on the day of the Wiki Freeze as well as bringing transferable tattoo samples of ''Cura'' to the European Jamboree as a form of marketing for the video and concept as a whole. <br />
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The team also produced a film with Amy Congdon exploring ''Cura'' itself and its many applications. They decided to angle the film as a product launch in order to capture the imagination and bring Quanticare and ''Cura'' to the public as a realistic future involving synthetic biology. The film is available to view below, we thank the Biochemical Society for their outreach grant to fund this film.<br />
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<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table></center></html><br />
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<html><font size=3pt><b>Please check out <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details!</b></font></html><br />
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Some initial feedback, the Quanticare video was launched less than 24 hours before the wiki freeze:<br />
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-Blogged about on Biochemical Society blog<br />
-On UEA news pages <br />
-Mentioned in tweet from the BBSRC<br />
-Many likes and tweets<br />
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-Some quotes <br />
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"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
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"Great project. Good luck!" @SocratesLogos Synthetic Biologist<br />
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"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
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==Social Media==<br />
The NRPUEA iGEM team has been VERY active on a range of social media over the summer.Any one is welcome to follow the team on Facebook or twitter. The team kept everyone updated on what was going on, both in the lab and in our spare time. This has been a great method of connecting with the public, companies,researchers, as well as getting in touch with other iGEM teams. The NRPUEA iGEM team believe human out reach via social media is important since it allows the public to follow the teams achievements and lab results, as well as providing an opportunity for members of the public to get to know the personality of the team members. The human stories behind science are crucial, informative and a powerful way to promote science in a positive light. Please follow us on twitter @NRPiGEM or UEA iGEM 2012 page on Facebook.<br />
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===Twitter===<br />
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[[File:Twitter stats NRP iGEM.png]]<br />
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[[File:TweetReach_nrpigem.pdf| '''Download a PDF of our twitter statistics''']]<br />
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===Facebook===<br />
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Creating social media pages also allowed us to access information about who was checking our page out, showing interesting information, showing that social media techniques such as facebook attracted mostly 18-24 years olds interest, and that it attracted no attention from those above 65. Therefore the team thought it was important to promote our project, not only through the internet, but also other techniques such as newspaper articles and radio shows. <br />
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The figure bellow shows the age ranges and gender of people who liked any posts from the NRP UEA iGEM team<br />
[[File:Likes.png |600px| centre]]<br />
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Below is a list of the international locations that people liked our teams status updates from. This highlighted that whilst the majority of followers were located in Norwich, we also had followers from a range of places over the world, highlighting the importance of using such social media to reach these followers. <br />
[[File:Location_of_likes.png |600px| centre]]<br />
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As mentioned the NRP UEA iGEM team created facebook statuses and tweets very regularly and as the graph below shows, there was a consistent increase in action and interest in the NRPUEA iGEM team throughout the summer. <br />
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[[File:Posts_ratings.png |600px| centre]]<br />
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===Google+===<br />
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We championed the use of Google+, a new social network from Google. We encouraged the UK iGEM teams to utilise Google+ to help make the 2012 UK team meetup as open and accessible as possible.<br />
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==Media appearances==<br />
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===STAR Radio===<br />
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[http://www.star107.co.uk/farming-show.php STAR 107.9 FM's Farming Show]<br />
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[[Image:NRPSTARRadio.jpg | 300px | right]]<br />
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[http://www.star107.co.uk/podcasts/the-farming-show/show-23.mp3 Listen To Our Radio Show Appearence!]<br />
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In early July the team travelled to Cambridge to appear on STAR Radio's program: "The Farming Show". We were interviewed by Mark Peters (one of the presenters) about synthetic biology, iGEM and our project. The interview went really well as Pascoe and Khadija got across all of the information that was needed in a clear and concise way, and the presenter was extremely helpful in making sure all of the salient points of the project were brought out. <br />
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The team were able to give a brief description of what synthetic biology is before talking about the future applications of synthetic biology for medicine, agriculture and business. We were also able to mention our event at the Norwich Forum in late August in order to help promote it!<br />
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A big thanks goes out to all at STAR Radio 107.9 for their help with the project in setting up the interview and allowing us air time to discuss iGEM and synthetic biology. Please visit the link above to hear the radio show.<br />
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===Newspapers & Internet Promotion===<br />
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[[File:Paper.jpg|200px|left]]<br />
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The team was featured on the UEA website throughout the summer including a press release regarding the entire project ([http://www.uea.ac.uk/mac/comm/media/press/2012/August/igem-competition-uea Found Here]) and a press release regarding the Quanticare/''Cura'' future of synthetic biology video ([http://www.uea.ac.uk/bio/news/fb26092012?utm_source=info&utm_medium=email&utm_term=sept+26&utm_content=bio+staff&utm_campaign=comms Found Here]). We are extremely grateful to the UEA Press Office for their help in publicising our project and the events/outreach material we have produced as part of it.<br />
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We have also been featured in two local newspapers; the Norwich Evening News (pictured left) and the Eastern Daily Press. This has allowed us to further engage the public with our work and help show the project to the local community. We hope that these articles have helped to promote synthetic biology and introduce it to members of the public who would otherwise have not heard about it. We thank the teams at both newspapers for covering the story of our project.<br />
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Further to this a UEA graduate, Daniel Vipond, has followed our project and iGEM as a whole in order to produce a report for his journalism career. It has been extremely exciting to be followed by a television camera throughout the last few weeks and hope Daniel does very well with the report!<br />
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==Engagement with the wider scientific community==<br />
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=== The team presented at the John Innes Centre Synthetic Biology conference ===<br />
[[File:JIC.png|200px|right]]<br />
The John Innes Center (JIC) is a world leading center of plant research, the home of UEAs 2011 team, and part of the Norwich Research Park (NRP). Researchers at the JIC recognized that synthetic biology was an important field and therefore hosted a synthetic biology conference. The aim of the day was to enhance local researchers' knowledge of what synthetic biology is, and highlight projects in which synthetic biology is currently being used. When the team was invited to give a talk to the JIC researchers we were extremely excited. At the talk we gave an over view of what iGEM is, our project and how it developed over the summer, as well informing them about aspects of human practice. The team had carried out over the summer. the team took also showed them a clip of our futuristic applications film. We enjoyed presenting our work and results, as well having an opportunity to improve our presentation skills before the European jamboree. There seemed to be a real interest by the researchers in to our project, and the advice given was encouraging giving encouraging advice. The researchers also seemed very interested to hear more about our human practices elements of iGEM, wanting to know what we had discovered about the public's opinion our research. More than anything, researchers at the John Innes Centre couldn't believe that we were undergraduate students who had been working together for 10 weeks!<br />
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The NRP iGEM teams are helping to foster collaborations between scientists across the NRP and have begun to create a sense of community amongst NRP synthetic biologists.<br />
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===SGM microbiology synthetic biology (September 3rd -5th)===<br />
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The society of general microbiology held a 3 day synthetic biology conference at the University of Warwick. Three members of the UEA NRP iGEM team were lucky enough to go and enjoy talks on streptococcus, molecular motors, and the dynamics of the genome and designer microbes. The event also gave the team member the opportunity to meet other synthetic biology scientists as well as informing them of the current advances in the field and inspiring them with the future possibilities. The event also allowed the team members to present a poster and therefore engage with the scientific community, both informing them more about the iGEM competition and our project . The team believed it was important for the scientists in the synthetic biology world to have the opportunity to meet undergraduate iGEM members, allowing them to express the skills it has allowed them to develop and to encourage further interest in to lab groups forming teams, and funding iGEM projects.<br />
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=== The Biochemical Society kindly mentioned us in their blog post ===<br />
<br />
Not only did the Biochemical Society generously fund our project, they also were kind enough to feature our teams efforts in an encouraging ethical debate around the subject of synthetic biology (via our Quanticare video) in their blog.<br />
<br />
[http://biochemicalsociety.wordpress.com/2012/09/26/exploring-ethical-dilemmas-of-synthetic-biology-with-the-nrp-uea-norwich-igem-team/ '''Click here to view the blog post''']<br />
<br />
This post was also seen by the BBSRC, who very kindly tweeted about it!<br />
<br />
[[File:BBSRC.png | centre | 600px]]</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/Human_OutreachTeam:NRP-UEA-Norwich/Human Outreach2012-09-27T00:33:18Z<p>RussellGritton: /* Newspapers & Internet Promotion */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Human Practices=<br />
<br />
<br />
The UEA NRP team really went up and beyond this year, carrying out a wide selection of activity's to inform the public of both synthetic biology and iGEM, aiming to raise awareness of this branch of science. We used many methods of communication in order to cater for a range of audiences, including a radio show, newspaper articles, pubic events and talks as well as collaborating with all the other UK teams, in order to organize the UK team meet up and steam it live on the internet. <br />
<br />
== iGEM collaborations==<br />
<br />
===The iGEM UK team meet up, hosted by the NRP-UEA team at Google campus London (Friday 17th August) The most open and publically accessible UK team meetup EVER===<br />
<br />
[[File:Hangout stats.png | center]]<br />
<br />
[[File:UKiGEMteams.jpg | 300px | right]]<br />
<br />
The NRPUEA iGEM team organised and hosted the UK team meet up at Google campus in London. The NRPUEA iGEM team greeted the UK teams, provided refreshments and a lovely buffet lunch, as well as chairing the speeches and making sure the day ran smoothly. There was a great atmosphere at the meet up, and the event was a huge success.<br />
<br />
The day consisted of a number of guest speakers, including advice and tips for iGEM success from Dr. Tom Ellis (advisor of the 2009 winning iGEM project E.chromi) and an interactive talk on synthetic biology and science communication from the BBC science presenter, Nature editor and Guardian writer, Dr. Adam Rutherford. The UK iGEM teams presented a poster and a 15 minute presentation about their project. This gave the members of each team the chance to practice their presentation skills before the European team meet up, as well as a chance for others to ask questions and see what other project routes the other teams have taken. <br />
<br />
The event was also available '''LIVE on the internet''' via Google+ hangout, to allow the public and those that were unable to attend to still watch the presentations. Despite there being a few technical difficulties with the quality of the image, figures have shown that over 600 people watched the event.<br />
<br />
[https://dl.dropbox.com/u/9957127/UK%20iGEM%20Team%20Hangout%2017th%20Aug%202012%20Programme.pdf '''Click here for programme PDF''']<br />
<br />
<br><br />
<br />
<html><align=center><br />
<table align=center width=100% cellpadding=0 cellspacing=0><br />
<tr><br />
<td valign=absmiddle align=center><br />
<iframe width="560" height="315" src="http://www.youtube.com/embed/Kv_Z14OIrKc" frameborder="0" allowfullscreen></iframe><br />
</td><br />
</tr><br />
</table><br />
</align><br />
</html><br />
<br />
<br><br />
<br />
=== Communicating with other iGEM teams===<br />
<br />
As part of our efforts to collaborate with the other iGEM teams this year we've made an effort to keep in contact with an array of iGEM teams. Here are some highlights:<br />
<br />
- We communicated with all of the UK iGEM teams via Twitter, Email, Facebook and Google+ before, during and after the UK team meet up.<br />
<br />
- We tweeted with many other iGEM teams from around the world and closely followed the iGEM community.<br />
<br />
- We video called the St Andrews iGEM team via Google+ Hang-out to discuss our projects further as well as to seek advice about the arrangements for the UK team meet-up.<br />
<br />
- Our advisor Richard Kelwick was interviewed by the Edinburgh iGEM team via Skype about our teams perspectives on public awareness and understanding of synthetic biology.<br />
<br />
- We visited the Cambridge team near the beginning of our project and they were kind enough to give us a tour of their labs. <br />
<br />
- Finally, the British Columbia team sent us a questionnaire about intellectual property that we happily filled out.<br />
<br />
==Public engagement==<br />
<br />
===The Future of Science event held at the forum, Norwich (Sunday 19th August)===<br />
<br />
<br />
<br />
[[File:Forum_team.JPG | 300px| right]]<br />
<br />
<br />
The Forum is located at the heart of the city of Norwich, and hosts a wide range of free events and exhibits to be enjoyed by the whole community. The venue is packed with restaurants, cafes and shops, as well as being home to Norwich library and BBC East Anglia. <br />
<br />
[[File:Poster for Forum.jpg | 250px | left]]<br />
<br />
The NRPUEA iGEM team were fortunate enough to hold an interactive day at the Forum, giving the public a chance to engage within the world of synthetic biology. Before the date of the event, the team increased awareness via a range of techniques, including posters, leaflets and newspaper and internet adverts, including the Forums website. This resulted with a large turnout at the event, including the local press. <br />
<br />
At the Future of Science event, the team presented a selection of informative posters, revealed their new film on a large screen, as well as showing examples of work carried out by the students during iGEM. For example, the public could view ''E.coli'' which had been transformed with GFP within a UV light box, as well as carrying out a counting colonies exercise. The NRPUEA team also created a range of synthetic biology educational material for the event, to inform the public of the concepts and terminology of synthetic biology. The different activities were designed to interest a selection of different ages. The educational resources include a range of quizzes and worksheets such as crosswords and matching exercises.<br />
<br />
[[File:Child_making_plasmid.jpg |300px | right]]<br />
<br />
For younger children we designed a colourful jigsaw plasmid (as seen in image). The idea of this activity was for the participants to choose jigsaw pieces that represented the genes that code for certain characteristics, such as green or red hair, and piece them together to form a complete plasmid. Once they had designed their plasmid they used a biscuit base, icing of different colours and various sweets to create a "biscuit creature" that corresponded to the plasmid they had created. The learning outcome of this activity was to give the children an understanding that a different selection of genes, results in different characteristics, and that editing the DNA can result in a change in the characteristics. The activity was very popular and most of the participants seemed to understand this concept, as well as having great fun.<br />
<br />
The event attracted a range of ages, as well as mixed levels of previous interest into synthetic biology. It also became clear that many members of the public knew very little about synthetic biology, and therefore were very interested in finding out more about the new branch of biology and the iGEM competition. The NRPUEA iGEM team also took the opportunity to talk to the public and listen to their opinions about the work the iGEM teams are carrying out, which demonstrated how mixed the view of the public were. The day was a huge success.<br />
<br />
'''Synthetic Biology activity sheets'''<br />
<br />
[[File:Word_Search_of_Synthetic_Biology.pdf]],<br />
[[File:Decode_Disney.pdf]],<br />
[[File:Questions_comprehensive.pdf]],[[File:Decode_Music_Artists_and_Films.pdf]],[[File:Word_Search_of_Synthetic_Biology.pdf]]<br />
<br />
=== Creation of a futuristic company, Quanticare and production of a thought-provoking film===<br />
<br />
<br />
[[File:Tattoo1.png |300px]][[File:QuanticareLogo_(2).png |300px]][[File:Tattoo2.png |300px]]<br />
<br />
<br />
As part of our human practices we decided to look into an artistic approach to bringing synthetic biology to a wider audience. As part of this we created "Quanticare", a fictitious and futuristic company forming on the back of a rise to prominence of synthetic biology over the next few years, as well as the results of our own projects on nitric oxide sensing and the comparator circuit. The fictitious company's latest development is to introduce a visual bio sensors in to the human body in the form of a tattoo-based health monitor, ''Cura''. <br />
<br />
<br />
Not only did the development of the fictitious company allow the team to explore potential future applications of synthetic biology and bio sensors, but also via the creation of a film , fuel a range of ethical questions. The team released the video on the day of the Wiki Freeze as well as bringing transferable tattoo samples of ''Cura'' to the European Jamboree as a form of marketing for the video and concept as a whole. <br />
<br />
<br />
The team also produced a film with Amy Congdon exploring ''Cura'' itself and its many applications. They decided to angle the film as a product launch in order to capture the imagination and bring Quanticare and ''Cura'' to the public as a realistic future involving synthetic biology. The film is available to view below, we thank the Biochemical Society for their outreach grant to fund this film.<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table></center></html><br />
<br />
<br />
<html><font size=3pt><b>Please check out <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details!</b></font></html><br />
<br />
<br><br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 hours before the wiki freeze:<br />
<br />
-Blogged about on Biochemical Society blog<br />
-On UEA news pages <br />
-Mentioned in tweet from the BBSRC<br />
-Many likes and tweets<br />
<br />
-Some quotes <br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
==Social Media==<br />
The NRPUEA iGEM team has been VERY active on a range of social media over the summer.Any one is welcome to follow the team on Facebook or twitter. The team kept everyone updated on what was going on, both in the lab and in our spare time. This has been a great method of connecting with the public, companies,researchers, as well as getting in touch with other iGEM teams. The NRPUEA iGEM team believe human out reach via social media is important since it allows the public to follow the teams achievements and lab results, as well as providing an opportunity for members of the public to get to know the personality of the team members. The human stories behind science are crucial, informative and a powerful way to promote science in a positive light. Please follow us on twitter @NRPiGEM or UEA iGEM 2012 page on Facebook.<br />
<br />
===Twitter===<br />
<br />
[[File:Twitter stats NRP iGEM.png]]<br />
<br />
[[File:TweetReach_nrpigem.pdf| '''Download a PDF of our twitter statistics''']]<br />
<br />
===Facebook===<br />
<br />
Creating social media pages also allowed us to access information about who was checking our page out, showing interesting information, showing that social media techniques such as facebook attracted mostly 18-24 years olds interest, and that it attracted no attention from those above 65. Therefore the team thought it was important to promote our project, not only through the internet, but also other techniques such as newspaper articles and radio shows. <br />
<br />
The figure bellow shows the age ranges and gender of people who liked any posts from the NRP UEA iGEM team<br />
[[File:Likes.png |600px| centre]]<br />
<br />
Below is a list of the international locations that people liked our teams status updates from. This highlighted that whilst the majority of followers were located in Norwich, we also had followers from a range of places over the world, highlighting the importance of using such social media to reach these followers. <br />
[[File:Location_of_likes.png |600px| centre]]<br />
<br />
As mentioned the NRP UEA iGEM team created facebook statuses and tweets very regularly and as the graph below shows, there was a consistent increase in action and interest in the NRPUEA iGEM team throughout the summer. <br />
<br />
[[File:Posts_ratings.png |600px| centre]]<br />
<br />
===Google+===<br />
<br />
We championed the use of Google+, a new social network from Google. We encouraged the UK iGEM teams to utilise Google+ to help make the 2012 UK team meetup as open and accessible as possible.<br />
<br />
==Media appearances==<br />
<br />
===STAR Radio===<br />
<br />
[http://www.star107.co.uk/farming-show.php STAR 107.9 FM's Farming Show]<br />
<br />
[[Image:NRPSTARRadio.jpg | 300px | right]]<br />
<br />
[http://www.star107.co.uk/podcasts/the-farming-show/show-23.mp3 Listen To Our Radio Show Appearence!]<br />
<br />
In early July the team travelled to Cambridge to appear on STAR Radio's program: "The Farming Show". We were interviewed by Mark Peters (one of the presenters) about synthetic biology, iGEM and our project. The interview went really well as Pascoe and Khadija got across all of the information that was needed in a clear and concise way, and the presenter was extremely helpful in making sure all of the salient points of the project were brought out. <br />
<br />
The team were able to give a brief description of what synthetic biology is before talking about the future applications of synthetic biology for medicine, agriculture and business. We were also able to mention our event at the Norwich Forum in late August in order to help promote it!<br />
<br />
A big thanks goes out to all at STAR Radio 107.9 for their help with the project in setting up the interview and allowing us air time to discuss iGEM and synthetic biology. Please visit the link above to hear the radio show.<br />
<br />
===Newspapers & Internet Promotion===<br />
<br />
[[File:Paper.jpg|200px|left]]<br />
<br />
The team was featured on the UEA website throughout the summer including a press release regarding the entire project ([http://www.uea.ac.uk/mac/comm/media/press/2012/August/igem-competition-uea Found Here]) and a press release regarding the Quanticare/''Cura'' future of synthetic biology video ([http://www.uea.ac.uk/bio/news/fb26092012?utm_source=info&utm_medium=email&utm_term=sept+26&utm_content=bio+staff&utm_campaign=comms Found Here]). We are extremely grateful to the UEA Press Office for their help in publicising our project and the events/outreach material we have produced as part of it.<br />
<br />
<br />
We have also been featured in two local newspapers; the Norwich Evening News (pictured left) and the Eastern Daily Press. This has allowed us to further engage the public with our work and help show the project to the local community. We hope that these articles have helped to promote synthetic biology and introduce it to members of the public who would otherwise have not heard about it. We thank the teams at both newspapers for covering the story of our project.<br />
<br />
<br />
Further to this a UEA graduate, Daniel Vipond, has followed our project and iGEM as a whole in order to produce a report for his journalism career. It has been extremely exciting to be followed by a television camera throughout the last few weeks and hope Daniel does very well with the report!<br />
<br><br><br />
<br />
==Engagement with the wider scientific community==<br />
<br />
=== The team presented at the John Innes Centre Synthetic Biology conference ===<br />
[[File:JIC.png|200px|right]]<br />
The John Innes Center (JIC) is a world leading center of plant research, the home of UEAs 2011 team, and part of the Norwich Research Park (NRP). Researchers at the JIC recognized that synthetic biology was an important field and therefore hosted a synthetic biology conference. The aim of the day was to enhance local researchers' knowledge of what synthetic biology is, and highlight projects in which synthetic biology is currently being used. When the team was invited to give a talk to the JIC researchers we were extremely excited. At the talk we gave an over view of what iGEM is, our project and how it developed over the summer, as well informing them about aspects of human practice. The team had carried out over the summer. the team took also showed them a clip of our futuristic applications film. We enjoyed presenting our work and results, as well having an opportunity to improve our presentation skills before the European jamboree. There seemed to be a real interest by the researchers in to our project, and the advice given was encouraging giving encouraging advice. The researchers also seemed very interested to hear more about our human practices elements of iGEM, wanting to know what we had discovered about the public's opinion our research. More than anything, researchers at the John Innes Centre couldn't believe that we were undergraduate students who had been working together for 10 weeks!<br />
<br />
The NRP iGEM teams are helping to foster collaborations between scientists across the NRP and have begun to create a sense of community amongst NRP synthetic biologists.<br />
<br />
===SGM microbiology synthetic biology (September 3rd -5th)===<br />
<br />
The society of general microbiology held a 3 day synthetic biology conference at the University of Warwick. Three members of the UEA NRP iGEM team were lucky enough to go and enjoy talks on streptococcus, molecular motors, and the dynamics of the genome and designer microbes. The event also gave the team member the opportunity to meet other synthetic biology scientists as well as informing them of the current advances in the field and inspiring them with the future possibilities. The event also allowed the team members to present a poster and therefore engage with the scientific community, both informing them more about the iGEM competition and our project . The team believed it was important for the scientists in the synthetic biology world to have the opportunity to meet undergraduate iGEM members, allowing them to express the skills it has allowed them to develop and to encourage further interest in to lab groups forming teams, and funding iGEM projects.<br />
<br />
=== The Biochemical Society kindly mentioned us in their blog post ===<br />
<br />
Not only did the Biochemical Society generously fund our project, they also were kind enough to feature our teams efforts in an encouraging ethical debate around the subject of synthetic biology (via our Quanticare video) in their blog.<br />
<br />
[http://biochemicalsociety.wordpress.com/2012/09/26/exploring-ethical-dilemmas-of-synthetic-biology-with-the-nrp-uea-norwich-igem-team/ '''Click here to view the blog post''']<br />
<br />
This post was also seen by the BBSRC, who very kindly tweeted about it!<br />
<br />
[[File:BBSRC.png | centre | 600px]]</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/Human_OutreachTeam:NRP-UEA-Norwich/Human Outreach2012-09-27T00:32:36Z<p>RussellGritton: /* Newspapers */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Human Practices=<br />
<br />
<br />
The UEA NRP team really went up and beyond this year, carrying out a wide selection of activity's to inform the public of both synthetic biology and iGEM, aiming to raise awareness of this branch of science. We used many methods of communication in order to cater for a range of audiences, including a radio show, newspaper articles, pubic events and talks as well as collaborating with all the other UK teams, in order to organize the UK team meet up and steam it live on the internet. <br />
<br />
== iGEM collaborations==<br />
<br />
===The iGEM UK team meet up, hosted by the NRP-UEA team at Google campus London (Friday 17th August) The most open and publically accessible UK team meetup EVER===<br />
<br />
[[File:Hangout stats.png | center]]<br />
<br />
[[File:UKiGEMteams.jpg | 300px | right]]<br />
<br />
The NRPUEA iGEM team organised and hosted the UK team meet up at Google campus in London. The NRPUEA iGEM team greeted the UK teams, provided refreshments and a lovely buffet lunch, as well as chairing the speeches and making sure the day ran smoothly. There was a great atmosphere at the meet up, and the event was a huge success.<br />
<br />
The day consisted of a number of guest speakers, including advice and tips for iGEM success from Dr. Tom Ellis (advisor of the 2009 winning iGEM project E.chromi) and an interactive talk on synthetic biology and science communication from the BBC science presenter, Nature editor and Guardian writer, Dr. Adam Rutherford. The UK iGEM teams presented a poster and a 15 minute presentation about their project. This gave the members of each team the chance to practice their presentation skills before the European team meet up, as well as a chance for others to ask questions and see what other project routes the other teams have taken. <br />
<br />
The event was also available '''LIVE on the internet''' via Google+ hangout, to allow the public and those that were unable to attend to still watch the presentations. Despite there being a few technical difficulties with the quality of the image, figures have shown that over 600 people watched the event.<br />
<br />
[https://dl.dropbox.com/u/9957127/UK%20iGEM%20Team%20Hangout%2017th%20Aug%202012%20Programme.pdf '''Click here for programme PDF''']<br />
<br />
<br><br />
<br />
<html><align=center><br />
<table align=center width=100% cellpadding=0 cellspacing=0><br />
<tr><br />
<td valign=absmiddle align=center><br />
<iframe width="560" height="315" src="http://www.youtube.com/embed/Kv_Z14OIrKc" frameborder="0" allowfullscreen></iframe><br />
</td><br />
</tr><br />
</table><br />
</align><br />
</html><br />
<br />
<br><br />
<br />
=== Communicating with other iGEM teams===<br />
<br />
As part of our efforts to collaborate with the other iGEM teams this year we've made an effort to keep in contact with an array of iGEM teams. Here are some highlights:<br />
<br />
- We communicated with all of the UK iGEM teams via Twitter, Email, Facebook and Google+ before, during and after the UK team meet up.<br />
<br />
- We tweeted with many other iGEM teams from around the world and closely followed the iGEM community.<br />
<br />
- We video called the St Andrews iGEM team via Google+ Hang-out to discuss our projects further as well as to seek advice about the arrangements for the UK team meet-up.<br />
<br />
- Our advisor Richard Kelwick was interviewed by the Edinburgh iGEM team via Skype about our teams perspectives on public awareness and understanding of synthetic biology.<br />
<br />
- We visited the Cambridge team near the beginning of our project and they were kind enough to give us a tour of their labs. <br />
<br />
- Finally, the British Columbia team sent us a questionnaire about intellectual property that we happily filled out.<br />
<br />
==Public engagement==<br />
<br />
===The Future of Science event held at the forum, Norwich (Sunday 19th August)===<br />
<br />
<br />
<br />
[[File:Forum_team.JPG | 300px| right]]<br />
<br />
<br />
The Forum is located at the heart of the city of Norwich, and hosts a wide range of free events and exhibits to be enjoyed by the whole community. The venue is packed with restaurants, cafes and shops, as well as being home to Norwich library and BBC East Anglia. <br />
<br />
[[File:Poster for Forum.jpg | 250px | left]]<br />
<br />
The NRPUEA iGEM team were fortunate enough to hold an interactive day at the Forum, giving the public a chance to engage within the world of synthetic biology. Before the date of the event, the team increased awareness via a range of techniques, including posters, leaflets and newspaper and internet adverts, including the Forums website. This resulted with a large turnout at the event, including the local press. <br />
<br />
At the Future of Science event, the team presented a selection of informative posters, revealed their new film on a large screen, as well as showing examples of work carried out by the students during iGEM. For example, the public could view ''E.coli'' which had been transformed with GFP within a UV light box, as well as carrying out a counting colonies exercise. The NRPUEA team also created a range of synthetic biology educational material for the event, to inform the public of the concepts and terminology of synthetic biology. The different activities were designed to interest a selection of different ages. The educational resources include a range of quizzes and worksheets such as crosswords and matching exercises.<br />
<br />
[[File:Child_making_plasmid.jpg |300px | right]]<br />
<br />
For younger children we designed a colourful jigsaw plasmid (as seen in image). The idea of this activity was for the participants to choose jigsaw pieces that represented the genes that code for certain characteristics, such as green or red hair, and piece them together to form a complete plasmid. Once they had designed their plasmid they used a biscuit base, icing of different colours and various sweets to create a "biscuit creature" that corresponded to the plasmid they had created. The learning outcome of this activity was to give the children an understanding that a different selection of genes, results in different characteristics, and that editing the DNA can result in a change in the characteristics. The activity was very popular and most of the participants seemed to understand this concept, as well as having great fun.<br />
<br />
The event attracted a range of ages, as well as mixed levels of previous interest into synthetic biology. It also became clear that many members of the public knew very little about synthetic biology, and therefore were very interested in finding out more about the new branch of biology and the iGEM competition. The NRPUEA iGEM team also took the opportunity to talk to the public and listen to their opinions about the work the iGEM teams are carrying out, which demonstrated how mixed the view of the public were. The day was a huge success.<br />
<br />
'''Synthetic Biology activity sheets'''<br />
<br />
[[File:Word_Search_of_Synthetic_Biology.pdf]],<br />
[[File:Decode_Disney.pdf]],<br />
[[File:Questions_comprehensive.pdf]],[[File:Decode_Music_Artists_and_Films.pdf]],[[File:Word_Search_of_Synthetic_Biology.pdf]]<br />
<br />
=== Creation of a futuristic company, Quanticare and production of a thought-provoking film===<br />
<br />
<br />
[[File:Tattoo1.png |300px]][[File:QuanticareLogo_(2).png |300px]][[File:Tattoo2.png |300px]]<br />
<br />
<br />
As part of our human practices we decided to look into an artistic approach to bringing synthetic biology to a wider audience. As part of this we created "Quanticare", a fictitious and futuristic company forming on the back of a rise to prominence of synthetic biology over the next few years, as well as the results of our own projects on nitric oxide sensing and the comparator circuit. The fictitious company's latest development is to introduce a visual bio sensors in to the human body in the form of a tattoo-based health monitor, ''Cura''. <br />
<br />
<br />
Not only did the development of the fictitious company allow the team to explore potential future applications of synthetic biology and bio sensors, but also via the creation of a film , fuel a range of ethical questions. The team released the video on the day of the Wiki Freeze as well as bringing transferable tattoo samples of ''Cura'' to the European Jamboree as a form of marketing for the video and concept as a whole. <br />
<br />
<br />
The team also produced a film with Amy Congdon exploring ''Cura'' itself and its many applications. They decided to angle the film as a product launch in order to capture the imagination and bring Quanticare and ''Cura'' to the public as a realistic future involving synthetic biology. The film is available to view below, we thank the Biochemical Society for their outreach grant to fund this film.<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table></center></html><br />
<br />
<br />
<html><font size=3pt><b>Please check out <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details!</b></font></html><br />
<br />
<br><br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 hours before the wiki freeze:<br />
<br />
-Blogged about on Biochemical Society blog<br />
-On UEA news pages <br />
-Mentioned in tweet from the BBSRC<br />
-Many likes and tweets<br />
<br />
-Some quotes <br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
==Social Media==<br />
The NRPUEA iGEM team has been VERY active on a range of social media over the summer.Any one is welcome to follow the team on Facebook or twitter. The team kept everyone updated on what was going on, both in the lab and in our spare time. This has been a great method of connecting with the public, companies,researchers, as well as getting in touch with other iGEM teams. The NRPUEA iGEM team believe human out reach via social media is important since it allows the public to follow the teams achievements and lab results, as well as providing an opportunity for members of the public to get to know the personality of the team members. The human stories behind science are crucial, informative and a powerful way to promote science in a positive light. Please follow us on twitter @NRPiGEM or UEA iGEM 2012 page on Facebook.<br />
<br />
===Twitter===<br />
<br />
[[File:Twitter stats NRP iGEM.png]]<br />
<br />
[[File:TweetReach_nrpigem.pdf| '''Download a PDF of our twitter statistics''']]<br />
<br />
===Facebook===<br />
<br />
Creating social media pages also allowed us to access information about who was checking our page out, showing interesting information, showing that social media techniques such as facebook attracted mostly 18-24 years olds interest, and that it attracted no attention from those above 65. Therefore the team thought it was important to promote our project, not only through the internet, but also other techniques such as newspaper articles and radio shows. <br />
<br />
The figure bellow shows the age ranges and gender of people who liked any posts from the NRP UEA iGEM team<br />
[[File:Likes.png |600px| centre]]<br />
<br />
Below is a list of the international locations that people liked our teams status updates from. This highlighted that whilst the majority of followers were located in Norwich, we also had followers from a range of places over the world, highlighting the importance of using such social media to reach these followers. <br />
[[File:Location_of_likes.png |600px| centre]]<br />
<br />
As mentioned the NRP UEA iGEM team created facebook statuses and tweets very regularly and as the graph below shows, there was a consistent increase in action and interest in the NRPUEA iGEM team throughout the summer. <br />
<br />
[[File:Posts_ratings.png |600px| centre]]<br />
<br />
===Google+===<br />
<br />
We championed the use of Google+, a new social network from Google. We encouraged the UK iGEM teams to utilise Google+ to help make the 2012 UK team meetup as open and accessible as possible.<br />
<br />
==Media appearances==<br />
<br />
===STAR Radio===<br />
<br />
[http://www.star107.co.uk/farming-show.php STAR 107.9 FM's Farming Show]<br />
<br />
[[Image:NRPSTARRadio.jpg | 300px | right]]<br />
<br />
[http://www.star107.co.uk/podcasts/the-farming-show/show-23.mp3 Listen To Our Radio Show Appearence!]<br />
<br />
In early July the team travelled to Cambridge to appear on STAR Radio's program: "The Farming Show". We were interviewed by Mark Peters (one of the presenters) about synthetic biology, iGEM and our project. The interview went really well as Pascoe and Khadija got across all of the information that was needed in a clear and concise way, and the presenter was extremely helpful in making sure all of the salient points of the project were brought out. <br />
<br />
The team were able to give a brief description of what synthetic biology is before talking about the future applications of synthetic biology for medicine, agriculture and business. We were also able to mention our event at the Norwich Forum in late August in order to help promote it!<br />
<br />
A big thanks goes out to all at STAR Radio 107.9 for their help with the project in setting up the interview and allowing us air time to discuss iGEM and synthetic biology. Please visit the link above to hear the radio show.<br />
<br />
===Newspapers & Internet Promotion===<br />
<br />
[[File:Paper.jpg|200px|left]]<br />
<br />
The team was featured on the UEA website throughout the summer including a press release regarding the entire project ([http://www.uea.ac.uk/mac/comm/media/press/2012/August/igem-competition-uea Found Here]) and a press release regarding the Quanticare/''Cura'' future of synthetic biology video ([http://www.uea.ac.uk/bio/news/fb26092012?utm_source=info&utm_medium=email&utm_term=sept+26&utm_content=bio+staff&utm_campaign=comms Found Here]). We are extremely grateful to the UEA Press Office for their help in publicising our project and the events/outreach material we have produced as part of it.<br />
<br />
<br />
We have also been featured in two local newspapers; the Norwich Evening News (pictured left) and the Eastern Daily Press. This has allowed us to further engage the public with our work and help show the project to the local community. We hope that these articles have helped to promote synthetic biology and introduce it to members of the public who would otherwise have not heard about it. We thank the teams at both newspapers for covering the story of our project.<br />
<br />
<br />
Further to this a UEA graduate, Daniel Vipond, has followed our project and iGEM as a whole in order to produce a report for his journalism career. It has been extremely exciting to be followed by a television camera throughout the last few weeks and hope Daniel does very well with the report!<br />
<br />
==Engagement with the wider scientific community==<br />
<br />
=== The team presented at the John Innes Centre Synthetic Biology conference ===<br />
[[File:JIC.png|200px|right]]<br />
The John Innes Center (JIC) is a world leading center of plant research, the home of UEAs 2011 team, and part of the Norwich Research Park (NRP). Researchers at the JIC recognized that synthetic biology was an important field and therefore hosted a synthetic biology conference. The aim of the day was to enhance local researchers' knowledge of what synthetic biology is, and highlight projects in which synthetic biology is currently being used. When the team was invited to give a talk to the JIC researchers we were extremely excited. At the talk we gave an over view of what iGEM is, our project and how it developed over the summer, as well informing them about aspects of human practice. The team had carried out over the summer. the team took also showed them a clip of our futuristic applications film. We enjoyed presenting our work and results, as well having an opportunity to improve our presentation skills before the European jamboree. There seemed to be a real interest by the researchers in to our project, and the advice given was encouraging giving encouraging advice. The researchers also seemed very interested to hear more about our human practices elements of iGEM, wanting to know what we had discovered about the public's opinion our research. More than anything, researchers at the John Innes Centre couldn't believe that we were undergraduate students who had been working together for 10 weeks!<br />
<br />
The NRP iGEM teams are helping to foster collaborations between scientists across the NRP and have begun to create a sense of community amongst NRP synthetic biologists.<br />
<br />
===SGM microbiology synthetic biology (September 3rd -5th)===<br />
<br />
The society of general microbiology held a 3 day synthetic biology conference at the University of Warwick. Three members of the UEA NRP iGEM team were lucky enough to go and enjoy talks on streptococcus, molecular motors, and the dynamics of the genome and designer microbes. The event also gave the team member the opportunity to meet other synthetic biology scientists as well as informing them of the current advances in the field and inspiring them with the future possibilities. The event also allowed the team members to present a poster and therefore engage with the scientific community, both informing them more about the iGEM competition and our project . The team believed it was important for the scientists in the synthetic biology world to have the opportunity to meet undergraduate iGEM members, allowing them to express the skills it has allowed them to develop and to encourage further interest in to lab groups forming teams, and funding iGEM projects.<br />
<br />
=== The Biochemical Society kindly mentioned us in their blog post ===<br />
<br />
Not only did the Biochemical Society generously fund our project, they also were kind enough to feature our teams efforts in an encouraging ethical debate around the subject of synthetic biology (via our Quanticare video) in their blog.<br />
<br />
[http://biochemicalsociety.wordpress.com/2012/09/26/exploring-ethical-dilemmas-of-synthetic-biology-with-the-nrp-uea-norwich-igem-team/ '''Click here to view the blog post''']<br />
<br />
This post was also seen by the BBSRC, who very kindly tweeted about it!<br />
<br />
[[File:BBSRC.png | centre | 600px]]</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/Human_OutreachTeam:NRP-UEA-Norwich/Human Outreach2012-09-27T00:26:21Z<p>RussellGritton: /* STAR Radio */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Human Practices=<br />
<br />
<br />
The UEA NRP team really went up and beyond this year, carrying out a wide selection of activity's to inform the public of both synthetic biology and iGEM, aiming to raise awareness of this branch of science. We used many methods of communication in order to cater for a range of audiences, including a radio show, newspaper articles, pubic events and talks as well as collaborating with all the other UK teams, in order to organize the UK team meet up and steam it live on the internet. <br />
<br />
== iGEM collaborations==<br />
<br />
===The iGEM UK team meet up, hosted by the NRP-UEA team at Google campus London (Friday 17th August) The most open and publically accessible UK team meetup EVER===<br />
<br />
[[File:Hangout stats.png | center]]<br />
<br />
[[File:UKiGEMteams.jpg | 300px | right]]<br />
<br />
The NRPUEA iGEM team organised and hosted the UK team meet up at Google campus in London. The NRPUEA iGEM team greeted the UK teams, provided refreshments and a lovely buffet lunch, as well as chairing the speeches and making sure the day ran smoothly. There was a great atmosphere at the meet up, and the event was a huge success.<br />
<br />
The day consisted of a number of guest speakers, including advice and tips for iGEM success from Dr. Tom Ellis (advisor of the 2009 winning iGEM project E.chromi) and an interactive talk on synthetic biology and science communication from the BBC science presenter, Nature editor and Guardian writer, Dr. Adam Rutherford. The UK iGEM teams presented a poster and a 15 minute presentation about their project. This gave the members of each team the chance to practice their presentation skills before the European team meet up, as well as a chance for others to ask questions and see what other project routes the other teams have taken. <br />
<br />
The event was also available '''LIVE on the internet''' via Google+ hangout, to allow the public and those that were unable to attend to still watch the presentations. Despite there being a few technical difficulties with the quality of the image, figures have shown that over 600 people watched the event.<br />
<br />
[https://dl.dropbox.com/u/9957127/UK%20iGEM%20Team%20Hangout%2017th%20Aug%202012%20Programme.pdf '''Click here for programme PDF''']<br />
<br />
<br><br />
<br />
<html><align=center><br />
<table align=center width=100% cellpadding=0 cellspacing=0><br />
<tr><br />
<td valign=absmiddle align=center><br />
<iframe width="560" height="315" src="http://www.youtube.com/embed/Kv_Z14OIrKc" frameborder="0" allowfullscreen></iframe><br />
</td><br />
</tr><br />
</table><br />
</align><br />
</html><br />
<br />
<br><br />
<br />
=== Communicating with other iGEM teams===<br />
<br />
As part of our efforts to collaborate with the other iGEM teams this year we've made an effort to keep in contact with an array of iGEM teams. Here are some highlights:<br />
<br />
- We communicated with all of the UK iGEM teams via Twitter, Email, Facebook and Google+ before, during and after the UK team meet up.<br />
<br />
- We tweeted with many other iGEM teams from around the world and closely followed the iGEM community.<br />
<br />
- We video called the St Andrews iGEM team via Google+ Hang-out to discuss our projects further as well as to seek advice about the arrangements for the UK team meet-up.<br />
<br />
- Our advisor Richard Kelwick was interviewed by the Edinburgh iGEM team via Skype about our teams perspectives on public awareness and understanding of synthetic biology.<br />
<br />
- We visited the Cambridge team near the beginning of our project and they were kind enough to give us a tour of their labs. <br />
<br />
- Finally, the British Columbia team sent us a questionnaire about intellectual property that we happily filled out.<br />
<br />
==Public engagement==<br />
<br />
===The Future of Science event held at the forum, Norwich (Sunday 19th August)===<br />
<br />
<br />
<br />
[[File:Forum_team.JPG | 300px| right]]<br />
<br />
<br />
The Forum is located at the heart of the city of Norwich, and hosts a wide range of free events and exhibits to be enjoyed by the whole community. The venue is packed with restaurants, cafes and shops, as well as being home to Norwich library and BBC East Anglia. <br />
<br />
[[File:Poster for Forum.jpg | 250px | left]]<br />
<br />
The NRPUEA iGEM team were fortunate enough to hold an interactive day at the Forum, giving the public a chance to engage within the world of synthetic biology. Before the date of the event, the team increased awareness via a range of techniques, including posters, leaflets and newspaper and internet adverts, including the Forums website. This resulted with a large turnout at the event, including the local press. <br />
<br />
At the Future of Science event, the team presented a selection of informative posters, revealed their new film on a large screen, as well as showing examples of work carried out by the students during iGEM. For example, the public could view ''E.coli'' which had been transformed with GFP within a UV light box, as well as carrying out a counting colonies exercise. The NRPUEA team also created a range of synthetic biology educational material for the event, to inform the public of the concepts and terminology of synthetic biology. The different activities were designed to interest a selection of different ages. The educational resources include a range of quizzes and worksheets such as crosswords and matching exercises.<br />
<br />
[[File:Child_making_plasmid.jpg |300px | right]]<br />
<br />
For younger children we designed a colourful jigsaw plasmid (as seen in image). The idea of this activity was for the participants to choose jigsaw pieces that represented the genes that code for certain characteristics, such as green or red hair, and piece them together to form a complete plasmid. Once they had designed their plasmid they used a biscuit base, icing of different colours and various sweets to create a "biscuit creature" that corresponded to the plasmid they had created. The learning outcome of this activity was to give the children an understanding that a different selection of genes, results in different characteristics, and that editing the DNA can result in a change in the characteristics. The activity was very popular and most of the participants seemed to understand this concept, as well as having great fun.<br />
<br />
The event attracted a range of ages, as well as mixed levels of previous interest into synthetic biology. It also became clear that many members of the public knew very little about synthetic biology, and therefore were very interested in finding out more about the new branch of biology and the iGEM competition. The NRPUEA iGEM team also took the opportunity to talk to the public and listen to their opinions about the work the iGEM teams are carrying out, which demonstrated how mixed the view of the public were. The day was a huge success.<br />
<br />
'''Synthetic Biology activity sheets'''<br />
<br />
[[File:Word_Search_of_Synthetic_Biology.pdf]],<br />
[[File:Decode_Disney.pdf]],<br />
[[File:Questions_comprehensive.pdf]],[[File:Decode_Music_Artists_and_Films.pdf]],[[File:Word_Search_of_Synthetic_Biology.pdf]]<br />
<br />
=== Creation of a futuristic company, Quanticare and production of a thought-provoking film===<br />
<br />
<br />
[[File:Tattoo1.png |300px]][[File:QuanticareLogo_(2).png |300px]][[File:Tattoo2.png |300px]]<br />
<br />
<br />
As part of our human practices we decided to look into an artistic approach to bringing synthetic biology to a wider audience. As part of this we created "Quanticare", a fictitious and futuristic company forming on the back of a rise to prominence of synthetic biology over the next few years, as well as the results of our own projects on nitric oxide sensing and the comparator circuit. The fictitious company's latest development is to introduce a visual bio sensors in to the human body in the form of a tattoo-based health monitor, ''Cura''. <br />
<br />
<br />
Not only did the development of the fictitious company allow the team to explore potential future applications of synthetic biology and bio sensors, but also via the creation of a film , fuel a range of ethical questions. The team released the video on the day of the Wiki Freeze as well as bringing transferable tattoo samples of ''Cura'' to the European Jamboree as a form of marketing for the video and concept as a whole. <br />
<br />
<br />
The team also produced a film with Amy Congdon exploring ''Cura'' itself and its many applications. They decided to angle the film as a product launch in order to capture the imagination and bring Quanticare and ''Cura'' to the public as a realistic future involving synthetic biology. The film is available to view below, we thank the Biochemical Society for their outreach grant to fund this film.<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table></center></html><br />
<br />
<br />
<html><font size=3pt><b>Please check out <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details!</b></font></html><br />
<br />
<br><br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 hours before the wiki freeze:<br />
<br />
-Blogged about on Biochemical Society blog<br />
-On UEA news pages <br />
-Mentioned in tweet from the BBSRC<br />
-Many likes and tweets<br />
<br />
-Some quotes <br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
==Social Media==<br />
The NRPUEA iGEM team has been VERY active on a range of social media over the summer.Any one is welcome to follow the team on Facebook or twitter. The team kept everyone updated on what was going on, both in the lab and in our spare time. This has been a great method of connecting with the public, companies,researchers, as well as getting in touch with other iGEM teams. The NRPUEA iGEM team believe human out reach via social media is important since it allows the public to follow the teams achievements and lab results, as well as providing an opportunity for members of the public to get to know the personality of the team members. The human stories behind science are crucial, informative and a powerful way to promote science in a positive light. Please follow us on twitter @NRPiGEM or UEA iGEM 2012 page on Facebook.<br />
<br />
===Twitter===<br />
<br />
[[File:Twitter stats NRP iGEM.png]]<br />
<br />
[[File:TweetReach_nrpigem.pdf| '''Download a PDF of our twitter statistics''']]<br />
<br />
===Facebook===<br />
<br />
Creating social media pages also allowed us to access information about who was checking our page out, showing interesting information, showing that social media techniques such as facebook attracted mostly 18-24 years olds interest, and that it attracted no attention from those above 65. Therefore the team thought it was important to promote our project, not only through the internet, but also other techniques such as newspaper articles and radio shows. <br />
<br />
The figure bellow shows the age ranges and gender of people who liked any posts from the NRP UEA iGEM team<br />
[[File:Likes.png |600px| centre]]<br />
<br />
Below is a list of the international locations that people liked our teams status updates from. This highlighted that whilst the majority of followers were located in Norwich, we also had followers from a range of places over the world, highlighting the importance of using such social media to reach these followers. <br />
[[File:Location_of_likes.png |600px| centre]]<br />
<br />
As mentioned the NRP UEA iGEM team created facebook statuses and tweets very regularly and as the graph below shows, there was a consistent increase in action and interest in the NRPUEA iGEM team throughout the summer. <br />
<br />
[[File:Posts_ratings.png |600px| centre]]<br />
<br />
===Google+===<br />
<br />
We championed the use of Google+, a new social network from Google. We encouraged the UK iGEM teams to utilise Google+ to help make the 2012 UK team meetup as open and accessible as possible.<br />
<br />
==Media appearances==<br />
<br />
===STAR Radio===<br />
<br />
[http://www.star107.co.uk/farming-show.php STAR 107.9 FM's Farming Show]<br />
<br />
[[Image:NRPSTARRadio.jpg | 300px | right]]<br />
<br />
[http://www.star107.co.uk/podcasts/the-farming-show/show-23.mp3 Listen To Our Radio Show Appearence!]<br />
<br />
In early July the team travelled to Cambridge to appear on STAR Radio's program: "The Farming Show". We were interviewed by Mark Peters (one of the presenters) about synthetic biology, iGEM and our project. The interview went really well as Pascoe and Khadija got across all of the information that was needed in a clear and concise way, and the presenter was extremely helpful in making sure all of the salient points of the project were brought out. <br />
<br />
The team were able to give a brief description of what synthetic biology is before talking about the future applications of synthetic biology for medicine, agriculture and business. We were also able to mention our event at the Norwich Forum in late August in order to help promote it!<br />
<br />
A big thanks goes out to all at STAR Radio 107.9 for their help with the project in setting up the interview and allowing us air time to discuss iGEM and synthetic biology. Please visit the link above to hear the radio show.<br />
<br />
===Newspapers===<br />
<br />
The team featured on the university website a few times through the summer, as well as a brief article in the Eastern daily press newspaper.This was great, as allowed the locals of Norwich to become aware not only of the team and iGEM, but also introduce many to the scientific branch of synthetic biology.<br />
<br />
<br />
<br />
<br />
<br />
==Engagement with the wider scientific community==<br />
<br />
=== The team presented at the John Innes Centre Synthetic Biology conference ===<br />
[[File:JIC.png|200px|right]]<br />
The John Innes Center (JIC) is a world leading center of plant research, the home of UEAs 2011 team, and part of the Norwich Research Park (NRP). Researchers at the JIC recognized that synthetic biology was an important field and therefore hosted a synthetic biology conference. The aim of the day was to enhance local researchers' knowledge of what synthetic biology is, and highlight projects in which synthetic biology is currently being used. When the team was invited to give a talk to the JIC researchers we were extremely excited. At the talk we gave an over view of what iGEM is, our project and how it developed over the summer, as well informing them about aspects of human practice. The team had carried out over the summer. the team took also showed them a clip of our futuristic applications film. We enjoyed presenting our work and results, as well having an opportunity to improve our presentation skills before the European jamboree. There seemed to be a real interest by the researchers in to our project, and the advice given was encouraging giving encouraging advice. The researchers also seemed very interested to hear more about our human practices elements of iGEM, wanting to know what we had discovered about the public's opinion our research. More than anything, researchers at the John Innes Centre couldn't believe that we were undergraduate students who had been working together for 10 weeks!<br />
<br />
The NRP iGEM teams are helping to foster collaborations between scientists across the NRP and have begun to create a sense of community amongst NRP synthetic biologists.<br />
<br />
===SGM microbiology synthetic biology (September 3rd -5th)===<br />
<br />
The society of general microbiology held a 3 day synthetic biology conference at the University of Warwick. Three members of the UEA NRP iGEM team were lucky enough to go and enjoy talks on streptococcus, molecular motors, and the dynamics of the genome and designer microbes. The event also gave the team member the opportunity to meet other synthetic biology scientists as well as informing them of the current advances in the field and inspiring them with the future possibilities. The event also allowed the team members to present a poster and therefore engage with the scientific community, both informing them more about the iGEM competition and our project . The team believed it was important for the scientists in the synthetic biology world to have the opportunity to meet undergraduate iGEM members, allowing them to express the skills it has allowed them to develop and to encourage further interest in to lab groups forming teams, and funding iGEM projects.<br />
<br />
=== The Biochemical Society kindly mentioned us in their blog post ===<br />
<br />
Not only did the Biochemical Society generously fund our project, they also were kind enough to feature our teams efforts in an encouraging ethical debate around the subject of synthetic biology (via our Quanticare video) in their blog.<br />
<br />
[http://biochemicalsociety.wordpress.com/2012/09/26/exploring-ethical-dilemmas-of-synthetic-biology-with-the-nrp-uea-norwich-igem-team/ '''Click here to view the blog post''']<br />
<br />
This post was also seen by the BBSRC, who very kindly tweeted about it!<br />
<br />
[[File:BBSRC.png | centre | 600px]]</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/Human_OutreachTeam:NRP-UEA-Norwich/Human Outreach2012-09-27T00:20:29Z<p>RussellGritton: /* The team presented at the John Innes Centre Synthetic Biology conference */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Human Practices=<br />
<br />
<br />
The UEA NRP team really went up and beyond this year, carrying out a wide selection of activity's to inform the public of both synthetic biology and iGEM, aiming to raise awareness of this branch of science. We used many methods of communication in order to cater for a range of audiences, including a radio show, newspaper articles, pubic events and talks as well as collaborating with all the other UK teams, in order to organize the UK team meet up and steam it live on the internet. <br />
<br />
== iGEM collaborations==<br />
<br />
===The iGEM UK team meet up, hosted by the NRP-UEA team at Google campus London (Friday 17th August) The most open and publically accessible UK team meetup EVER===<br />
<br />
[[File:Hangout stats.png | center]]<br />
<br />
[[File:UKiGEMteams.jpg | 300px | right]]<br />
<br />
The NRPUEA iGEM team organised and hosted the UK team meet up at Google campus in London. The NRPUEA iGEM team greeted the UK teams, provided refreshments and a lovely buffet lunch, as well as chairing the speeches and making sure the day ran smoothly. There was a great atmosphere at the meet up, and the event was a huge success.<br />
<br />
The day consisted of a number of guest speakers, including advice and tips for iGEM success from Dr. Tom Ellis (advisor of the 2009 winning iGEM project E.chromi) and an interactive talk on synthetic biology and science communication from the BBC science presenter, Nature editor and Guardian writer, Dr. Adam Rutherford. The UK iGEM teams presented a poster and a 15 minute presentation about their project. This gave the members of each team the chance to practice their presentation skills before the European team meet up, as well as a chance for others to ask questions and see what other project routes the other teams have taken. <br />
<br />
The event was also available '''LIVE on the internet''' via Google+ hangout, to allow the public and those that were unable to attend to still watch the presentations. Despite there being a few technical difficulties with the quality of the image, figures have shown that over 600 people watched the event.<br />
<br />
[https://dl.dropbox.com/u/9957127/UK%20iGEM%20Team%20Hangout%2017th%20Aug%202012%20Programme.pdf '''Click here for programme PDF''']<br />
<br />
<br><br />
<br />
<html><align=center><br />
<table align=center width=100% cellpadding=0 cellspacing=0><br />
<tr><br />
<td valign=absmiddle align=center><br />
<iframe width="560" height="315" src="http://www.youtube.com/embed/Kv_Z14OIrKc" frameborder="0" allowfullscreen></iframe><br />
</td><br />
</tr><br />
</table><br />
</align><br />
</html><br />
<br />
<br><br />
<br />
=== Communicating with other iGEM teams===<br />
<br />
As part of our efforts to collaborate with the other iGEM teams this year we've made an effort to keep in contact with an array of iGEM teams. Here are some highlights:<br />
<br />
- We communicated with all of the UK iGEM teams via Twitter, Email, Facebook and Google+ before, during and after the UK team meet up.<br />
<br />
- We tweeted with many other iGEM teams from around the world and closely followed the iGEM community.<br />
<br />
- We video called the St Andrews iGEM team via Google+ Hang-out to discuss our projects further as well as to seek advice about the arrangements for the UK team meet-up.<br />
<br />
- Our advisor Richard Kelwick was interviewed by the Edinburgh iGEM team via Skype about our teams perspectives on public awareness and understanding of synthetic biology.<br />
<br />
- We visited the Cambridge team near the beginning of our project and they were kind enough to give us a tour of their labs. <br />
<br />
- Finally, the British Columbia team sent us a questionnaire about intellectual property that we happily filled out.<br />
<br />
==Public engagement==<br />
<br />
===The Future of Science event held at the forum, Norwich (Sunday 19th August)===<br />
<br />
<br />
<br />
[[File:Forum_team.JPG | 300px| right]]<br />
<br />
<br />
The Forum is located at the heart of the city of Norwich, and hosts a wide range of free events and exhibits to be enjoyed by the whole community. The venue is packed with restaurants, cafes and shops, as well as being home to Norwich library and BBC East Anglia. <br />
<br />
[[File:Poster for Forum.jpg | 250px | left]]<br />
<br />
The NRPUEA iGEM team were fortunate enough to hold an interactive day at the Forum, giving the public a chance to engage within the world of synthetic biology. Before the date of the event, the team increased awareness via a range of techniques, including posters, leaflets and newspaper and internet adverts, including the Forums website. This resulted with a large turnout at the event, including the local press. <br />
<br />
At the Future of Science event, the team presented a selection of informative posters, revealed their new film on a large screen, as well as showing examples of work carried out by the students during iGEM. For example, the public could view ''E.coli'' which had been transformed with GFP within a UV light box, as well as carrying out a counting colonies exercise. The NRPUEA team also created a range of synthetic biology educational material for the event, to inform the public of the concepts and terminology of synthetic biology. The different activities were designed to interest a selection of different ages. The educational resources include a range of quizzes and worksheets such as crosswords and matching exercises.<br />
<br />
[[File:Child_making_plasmid.jpg |300px | right]]<br />
<br />
For younger children we designed a colourful jigsaw plasmid (as seen in image). The idea of this activity was for the participants to choose jigsaw pieces that represented the genes that code for certain characteristics, such as green or red hair, and piece them together to form a complete plasmid. Once they had designed their plasmid they used a biscuit base, icing of different colours and various sweets to create a "biscuit creature" that corresponded to the plasmid they had created. The learning outcome of this activity was to give the children an understanding that a different selection of genes, results in different characteristics, and that editing the DNA can result in a change in the characteristics. The activity was very popular and most of the participants seemed to understand this concept, as well as having great fun.<br />
<br />
The event attracted a range of ages, as well as mixed levels of previous interest into synthetic biology. It also became clear that many members of the public knew very little about synthetic biology, and therefore were very interested in finding out more about the new branch of biology and the iGEM competition. The NRPUEA iGEM team also took the opportunity to talk to the public and listen to their opinions about the work the iGEM teams are carrying out, which demonstrated how mixed the view of the public were. The day was a huge success.<br />
<br />
'''Synthetic Biology activity sheets'''<br />
<br />
[[File:Word_Search_of_Synthetic_Biology.pdf]],<br />
[[File:Decode_Disney.pdf]],<br />
[[File:Questions_comprehensive.pdf]],[[File:Decode_Music_Artists_and_Films.pdf]],[[File:Word_Search_of_Synthetic_Biology.pdf]]<br />
<br />
=== Creation of a futuristic company, Quanticare and production of a thought-provoking film===<br />
<br />
<br />
[[File:Tattoo1.png |300px]][[File:QuanticareLogo_(2).png |300px]][[File:Tattoo2.png |300px]]<br />
<br />
<br />
As part of our human practices we decided to look into an artistic approach to bringing synthetic biology to a wider audience. As part of this we created "Quanticare", a fictitious and futuristic company forming on the back of a rise to prominence of synthetic biology over the next few years, as well as the results of our own projects on nitric oxide sensing and the comparator circuit. The fictitious company's latest development is to introduce a visual bio sensors in to the human body in the form of a tattoo-based health monitor, ''Cura''. <br />
<br />
<br />
Not only did the development of the fictitious company allow the team to explore potential future applications of synthetic biology and bio sensors, but also via the creation of a film , fuel a range of ethical questions. The team released the video on the day of the Wiki Freeze as well as bringing transferable tattoo samples of ''Cura'' to the European Jamboree as a form of marketing for the video and concept as a whole. <br />
<br />
<br />
The team also produced a film with Amy Congdon exploring ''Cura'' itself and its many applications. They decided to angle the film as a product launch in order to capture the imagination and bring Quanticare and ''Cura'' to the public as a realistic future involving synthetic biology. The film is available to view below, we thank the Biochemical Society for their outreach grant to fund this film.<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table></center></html><br />
<br />
<br />
<html><font size=3pt><b>Please check out <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details!</b></font></html><br />
<br />
<br><br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 hours before the wiki freeze:<br />
<br />
-Blogged about on Biochemical Society blog<br />
-On UEA news pages <br />
-Mentioned in tweet from the BBSRC<br />
-Many likes and tweets<br />
<br />
-Some quotes <br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
==Social Media==<br />
The NRPUEA iGEM team has been VERY active on a range of social media over the summer.Any one is welcome to follow the team on Facebook or twitter. The team kept everyone updated on what was going on, both in the lab and in our spare time. This has been a great method of connecting with the public, companies,researchers, as well as getting in touch with other iGEM teams. The NRPUEA iGEM team believe human out reach via social media is important since it allows the public to follow the teams achievements and lab results, as well as providing an opportunity for members of the public to get to know the personality of the team members. The human stories behind science are crucial, informative and a powerful way to promote science in a positive light. Please follow us on twitter @NRPiGEM or UEA iGEM 2012 page on Facebook.<br />
<br />
===Twitter===<br />
<br />
[[File:Twitter stats NRP iGEM.png]]<br />
<br />
[[File:TweetReach_nrpigem.pdf| '''Download a PDF of our twitter statistics''']]<br />
<br />
===Facebook===<br />
<br />
Creating social media pages also allowed us to access information about who was checking our page out, showing interesting information, showing that social media techniques such as facebook attracted mostly 18-24 years olds interest, and that it attracted no attention from those above 65. Therefore the team thought it was important to promote our project, not only through the internet, but also other techniques such as newspaper articles and radio shows. <br />
<br />
The figure bellow shows the age ranges and gender of people who liked any posts from the NRP UEA iGEM team<br />
[[File:Likes.png |600px| centre]]<br />
<br />
Below is a list of the international locations that people liked our teams status updates from. This highlighted that whilst the majority of followers were located in Norwich, we also had followers from a range of places over the world, highlighting the importance of using such social media to reach these followers. <br />
[[File:Location_of_likes.png |600px| centre]]<br />
<br />
As mentioned the NRP UEA iGEM team created facebook statuses and tweets very regularly and as the graph below shows, there was a consistent increase in action and interest in the NRPUEA iGEM team throughout the summer. <br />
<br />
[[File:Posts_ratings.png |600px| centre]]<br />
<br />
===Google+===<br />
<br />
We championed the use of Google+, a new social network from Google. We encouraged the UK iGEM teams to utilise Google+ to help make the 2012 UK team meetup as open and accessible as possible.<br />
<br />
==Media appearances==<br />
<br />
===STAR Radio===<br />
<br />
[http://www.star107.co.uk/farming-show.php STAR 107.9 FM's Farming Show]<br />
<br />
[[Image:NRPSTARRadio.jpg | 300px | right]]<br />
<br />
[http://www.star107.co.uk/podcasts/the-farming-show/show-23.mp3 Listen To Our Radio Show Appearence!]<br />
<br />
In early July the team travelled to Cambridge to appear on STAR Radio's program: "The Farming Show". We were interviewed by Mark Peters (one of the presenters) about synthetic biology, iGEM and our project. The interview went really well as Pascoe and Khadija got across all of the information that was needed in a clear and concise way, and the presenter was extremely helpful in making sure all of the salient points of the project were brought out. <br />
<br />
The team were able to give a brief description of what synthetic biology is before talking about the future applications of synthetic biology for medicine, agriculture and business. We were also able to mention our event at the Norwich Forum in late August in order to help promote it!<br />
<br />
A big thanks goes out to all at STAR Radio 107.9 for their help with the project in setting up the interview and allowing us air time to discuss iGEM and synthetic biology. Please visit the link above to hear the radio show.<br />
<br />
<br />
<br />
[[File:Paper.jpg|100px|left]]<br />
<br />
===Newspapers===<br />
<br />
The team featured on the university website a few times through the summer, as well as a brief article in the Eastern daily press newspaper.This was great, as allowed the locals of Norwich to become aware not only of the team and iGEM, but also introduce many to the scientific branch of synthetic biology.<br />
<br />
<br />
<br />
<br />
<br />
==Engagement with the wider scientific community==<br />
<br />
=== The team presented at the John Innes Centre Synthetic Biology conference ===<br />
[[File:JIC.png|200px|right]]<br />
The John Innes Center (JIC) is a world leading center of plant research, the home of UEAs 2011 team, and part of the Norwich Research Park (NRP). Researchers at the JIC recognized that synthetic biology was an important field and therefore hosted a synthetic biology conference. The aim of the day was to enhance local researchers' knowledge of what synthetic biology is, and highlight projects in which synthetic biology is currently being used. When the team was invited to give a talk to the JIC researchers we were extremely excited. At the talk we gave an over view of what iGEM is, our project and how it developed over the summer, as well informing them about aspects of human practice. The team had carried out over the summer. the team took also showed them a clip of our futuristic applications film. We enjoyed presenting our work and results, as well having an opportunity to improve our presentation skills before the European jamboree. There seemed to be a real interest by the researchers in to our project, and the advice given was encouraging giving encouraging advice. The researchers also seemed very interested to hear more about our human practices elements of iGEM, wanting to know what we had discovered about the public's opinion our research. More than anything, researchers at the John Innes Centre couldn't believe that we were undergraduate students who had been working together for 10 weeks!<br />
<br />
The NRP iGEM teams are helping to foster collaborations between scientists across the NRP and have begun to create a sense of community amongst NRP synthetic biologists.<br />
<br />
===SGM microbiology synthetic biology (September 3rd -5th)===<br />
<br />
The society of general microbiology held a 3 day synthetic biology conference at the University of Warwick. Three members of the UEA NRP iGEM team were lucky enough to go and enjoy talks on streptococcus, molecular motors, and the dynamics of the genome and designer microbes. The event also gave the team member the opportunity to meet other synthetic biology scientists as well as informing them of the current advances in the field and inspiring them with the future possibilities. The event also allowed the team members to present a poster and therefore engage with the scientific community, both informing them more about the iGEM competition and our project . The team believed it was important for the scientists in the synthetic biology world to have the opportunity to meet undergraduate iGEM members, allowing them to express the skills it has allowed them to develop and to encourage further interest in to lab groups forming teams, and funding iGEM projects.<br />
<br />
=== The Biochemical Society kindly mentioned us in their blog post ===<br />
<br />
Not only did the Biochemical Society generously fund our project, they also were kind enough to feature our teams efforts in an encouraging ethical debate around the subject of synthetic biology (via our Quanticare video) in their blog.<br />
<br />
[http://biochemicalsociety.wordpress.com/2012/09/26/exploring-ethical-dilemmas-of-synthetic-biology-with-the-nrp-uea-norwich-igem-team/ '''Click here to view the blog post''']<br />
<br />
This post was also seen by the BBSRC, who very kindly tweeted about it!<br />
<br />
[[File:BBSRC.png | centre | 600px]]</div>RussellGrittonhttp://2012.igem.org/File:JIC.pngFile:JIC.png2012-09-27T00:19:17Z<p>RussellGritton: uploaded a new version of &quot;File:JIC.png&quot;</p>
<hr />
<div></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPQuanticareTemplate:UEANRPQuanticare2012-09-27T00:15:26Z<p>RussellGritton: </p>
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* html .dropmenudiv_a a{ /*IE only hack*/<br />
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<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
background-color: #8a3c3d;<br />
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</head><br />
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<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>Future Applications</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Practices</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
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<div class="ddcolortabsline">&nbsp;</div><br />
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<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
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</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPQuanticareTemplate:UEANRPQuanticare2012-09-27T00:14:02Z<p>RussellGritton: </p>
<hr />
<div>[[Image:QuanticareHeader.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
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<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
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}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/9/9f/QuanticareBG.png");<br />
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background-color:#a9c4cd;<br />
<br />
}<br />
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/*Browser History*/<br />
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a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
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#menubar.top-menu {<br />
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#menubar.slide-menu {<br />
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/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
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<br />
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/*Code to remove/overlay standard iGEM Header for blank canvas*/<br />
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a{<br />
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#footer-box {<br />
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<head><br />
<title>NRP UEA iGEM 2012</title><br />
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<!-- CSS for Drop Down Tabs Menu #1 --><br />
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<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
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<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
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<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
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<script type="text/javascript"><br />
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</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:12:38Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
#welcome a:visited{<br />
color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
background-position:center top;<br />
background-repeat:no-repeat;<br />
background-color:#009443;<br />
<br />
}<br />
<br />
/*Browser History*/<br />
a:link {ccolor:#20548f;}<br />
a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
a:active {color:#20548f;}<br />
<br />
#menubar.top-menu {<br />
position: absolute;<br />
top:0px;<br />
left:0px;<br />
}<br />
<br />
#menubar.slide-menu {<br />
position: absolute;<br />
top:0px;<br />
right:0px;<br />
}<br />
/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
color: #1D1D1B;<br />
}<br />
<br />
<br />
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#top-section{<br />
background-color: #009443;<br />
height: 100px;<br />
border: none;<br />
width: 975px;<br />
height:20px<br />
}<br />
<br />
.left-menu:hover {<br />
background-color: transparent;<br />
}<br />
<br />
.right-menu li a, .right-menu li a:hover {<br />
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}<br />
<br />
<br />
<br />
#menubar {<br />
width: auto;<br />
}<br />
<br />
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position: absolute;<br />
display: none;<br />
}<br />
<br />
#search-controls{<br />
display: none;<br />
}<br />
<br />
.firstHeading{<br />
display: none;<br />
}<br />
<br />
a{<br />
outline:none;<br />
}<br />
<br />
/*Page Controls(how wide the content section is within the window - background colour)*/ <br />
#content {<br />
width: 950x;<br />
border: none;<br />
font-family: helvetica, arial, sans-serif;<br />
color: #555555;<br />
}<br />
<br />
#footer-box {<br />
width:1075px;<br />
}<br />
</style><br />
<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
<br />
var tabdropdown={<br />
disappeardelay: 200, //set delay in miliseconds before menu disappears onmouseout<br />
disablemenuclick: false, //when user clicks on a menu item with a drop down menu, disable menu item's link?<br />
enableiframeshim: 1, //1 or 0, for true or false<br />
<br />
//No need to edit beyond here////////////////////////<br />
dropmenuobj: null, ie: document.all, firefox: document.getElementById&&!document.all, previousmenuitem:null,<br />
currentpageurl: window.location.href.replace("http://"+window.location.hostname, "").replace(/^\//, ""), //get current page url (minus hostname, ie: http://www.dynamicdrive.com/)<br />
<br />
getposOffset:function(what, offsettype){<br />
var totaloffset=(offsettype=="center")? what.offsetcenter : what.offsetTop;<br />
var parentEl=what.offsetParent;<br />
while (parentEl!=null){<br />
totaloffset=(offsettype=="center")? totaloffset+parentEl.offsetcenter : totaloffset+parentEl.offsetTop;<br />
parentEl=parentEl.offsetParent;<br />
}<br />
return totaloffset;<br />
},<br />
<br />
showhide:function(obj, e, obj2){ //obj refers to drop down menu, obj2 refers to tab menu item mouse is currently over<br />
if (this.ie || this.firefox)<br />
this.dropmenuobj.style.center=this.dropmenuobj.style.top="-500px"<br />
if (e.type=="click" && obj.visibility==hidden || e.type=="mouseover"){<br />
if (obj2.parentNode.className.indexOf("default")==-1) //if tab isn't a default selected one<br />
obj2.parentNode.className="selected"<br />
obj.visibility="visible"<br />
}<br />
else if (e.type=="click")<br />
obj.visibility="hidden"<br />
},<br />
<br />
iecompattest:function(){<br />
return (document.compatMode && document.compatMode!="BackCompat")? document.documentElement : document.body<br />
},<br />
<br />
clearbrowseredge:function(obj, whichedge){<br />
var edgeoffset=0<br />
if (whichedge=="rightedge"){<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollLeft+this.standardbody.clientWidth-15 : window.pageXOffset+window.innerWidth-15<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetWidth<br />
if (windowedge-this.dropmenuobj.x < this.dropmenuobj.contentmeasure) //move menu to the left?<br />
edgeoffset=this.dropmenuobj.contentmeasure-obj.offsetWidth<br />
}<br />
else{<br />
var topedge=this.ie && !window.opera? this.standardbody.scrollTop : window.pageYOffset<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollTop+this.standardbody.clientHeight-15 : window.pageYOffset+window.innerHeight-18<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetHeight<br />
if (windowedge-this.dropmenuobj.y < this.dropmenuobj.contentmeasure){ //move up?<br />
edgeoffset=this.dropmenuobj.contentmeasure+obj.offsetHeight<br />
if ((this.dropmenuobj.y-topedge)<this.dropmenuobj.contentmeasure) //up no good either?<br />
edgeoffset=this.dropmenuobj.y+obj.offsetHeight-topedge<br />
}<br />
this.dropmenuobj.firstlink.style.borderTopWidth=(edgeoffset==0)? 0 : "1px" //Add 1px top border to menu if dropping up<br />
}<br />
return edgeoffset<br />
},<br />
<br />
dropit:function(obj, e, dropmenuID){<br />
if (this.dropmenuobj!=null){ //hide previous menu<br />
this.dropmenuobj.style.visibility="hidden" //hide menu<br />
if (this.previousmenuitem!=null && this.previousmenuitem!=obj){<br />
if (this.previousmenuitem.parentNode.className.indexOf("default")==-1) //If the tab isn't a default selected one<br />
this.previousmenuitem.parentNode.className=""<br />
}<br />
}<br />
this.clearhidemenu()<br />
if (this.ie||this.firefox){<br />
obj.onmouseout=function(){tabdropdown.delayhidemenu(obj)}<br />
obj.onclick=function(){return !tabdropdown.disablemenuclick} //disable main menu item link onclick?<br />
this.dropmenuobj=document.getElementById(dropmenuID)<br />
this.dropmenuobj.onmouseover=function(){tabdropdown.clearhidemenu()}<br />
this.dropmenuobj.onmouseout=function(e){tabdropdown.dynamichide(e, obj)}<br />
this.dropmenuobj.onclick=function(){tabdropdown.delayhidemenu(obj)}<br />
this.showhide(this.dropmenuobj.style, e, obj)<br />
this.dropmenuobj.x=this.getposOffset(obj, "center")<br />
this.dropmenuobj.y=this.getposOffset(obj, "top")<br />
this.dropmenuobj.style.left=this.dropmenuobj.x-this.clearbrowseredge(obj, "rightedge")+"px"<br />
this.dropmenuobj.style.top=this.dropmenuobj.y-this.clearbrowseredge(obj, "bottomedge")+obj.offsetHeight+1+"px"<br />
this.previousmenuitem=obj //remember main menu item mouse moved out from (and into current menu item)<br />
this.positionshim() //call iframe shim function<br />
}<br />
},<br />
<br />
contains_firefox:function(a, b) {<br />
while (b.parentNode)<br />
if ((b = b.parentNode) == a)<br />
return true;<br />
return false;<br />
},<br />
<br />
dynamichide:function(e, obj2){ //obj2 refers to tab menu item mouse is currently over<br />
var evtobj=window.event? window.event : e<br />
if (this.ie&&!this.dropmenuobj.contains(evtobj.toElement))<br />
this.delayhidemenu(obj2)<br />
else if (this.firefox&&e.currentTarget!= evtobj.relatedTarget&& !this.contains_firefox(evtobj.currentTarget, evtobj.relatedTarget))<br />
this.delayhidemenu(obj2)<br />
},<br />
<br />
delayhidemenu:function(obj2){<br />
this.delayhide=setTimeout(function(){tabdropdown.dropmenuobj.style.visibility='hidden'; if (obj2.parentNode.className.indexOf('default')==-1) obj2.parentNode.className=''},this.disappeardelay) //hide menu<br />
},<br />
<br />
clearhidemenu:function(){<br />
if (this.delayhide!="undefined")<br />
clearTimeout(this.delayhide)<br />
},<br />
<br />
positionshim:function(){ //display iframe shim function<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined"){<br />
if (this.dropmenuobj.style.visibility=="visible"){<br />
this.shimobject.style.width=this.dropmenuobj.offsetWidth+"px"<br />
this.shimobject.style.height=this.dropmenuobj.offsetHeight+"px"<br />
this.shimobject.style.center=this.dropmenuobj.style.center<br />
this.shimobject.style.top=this.dropmenuobj.style.top<br />
}<br />
this.shimobject.style.display=(this.dropmenuobj.style.visibility=="visible")? "block" : "none"<br />
}<br />
},<br />
<br />
hideshim:function(){<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined")<br />
this.shimobject.style.display='none'<br />
},<br />
<br />
isSelected:function(menuurl){<br />
var menuurl=menuurl.replace("http://"+menuurl.hostname, "").replace(/^\//, "")<br />
return (tabdropdown.currentpageurl==menuurl)<br />
},<br />
<br />
init:function(menuid, dselected){<br />
this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
var menuitems=document.getElementById(menuid).getElementsByTagName("a")<br />
for (var i=0; i<menuitems.length; i++){<br />
if (menuitems[i].getAttribute("rel")){<br />
var relvalue=menuitems[i].getAttribute("rel")<br />
document.getElementById(relvalue).firstlink=document.getElementById(relvalue).getElementsByTagName("a")[0]<br />
menuitems[i].onmouseover=function(e){<br />
var event=typeof e!="undefined"? e : window.event<br />
tabdropdown.dropit(this, event, this.getAttribute("rel"))<br />
}<br />
}<br />
if (dselected=="auto" && typeof setalready=="undefined" && this.isSelected(menuitems[i].href)){<br />
menuitems[i].parentNode.className+=" selected default"<br />
var setalready=true<br />
}<br />
else if (parseInt(dselected)==i)<br />
menuitems[i].parentNode.className+=" selected default"<br />
}<br />
}<br />
<br />
}<br />
</script><br />
<style><br />
.ddcolortabs{<br />
padding: 0;<br />
width: 100%;<br />
background: transparent;<br />
voice-family: "\"}\"";<br />
voice-family: inherit;<br />
}<br />
<br />
.ddcolortabs ul{<br />
font: normal 16px Calibri, sans-serif;<br />
margin:0;<br />
padding:0;<br />
list-style:none;<br />
}<br />
<br />
.ddcolortabs li{<br />
display:inline;<br />
margin:0 2px 0 0;<br />
padding:0;<br />
text-transform:uppercase;<br />
}<br />
<br />
<br />
.ddcolortabs a{<br />
float: left;<br />
color: white;<br />
background: #009443 url(media/color_tabs_left.gif) no-repeat left top;<br />
margin:0 2px 0 0;<br />
padding:0 0 0px 0px;<br />
text-decoration:none;<br />
letter-spacing: 1px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:left;<br />
display:block;<br />
background: transparent url(media/color_tabs_right.gif) no-repeat right top;<br />
padding: 2px 4px 1px 3.5px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:none;<br />
}<br />
<br />
.ddcolortabs a:hover{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs a:hover span{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs .selected a, #ddcolortabs .selected a span{ /*currently selected tab*/<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabsline{<br />
clear: both;<br />
padding: 0;<br />
width: 100%;<br />
height: 8px;<br />
line-height: 8px;<br />
background: #009443;<br />
border-top: 1px solid #fff; /*Remove this to remove border between bar and tabs*/<br />
}<br />
<br />
/* ######### Style for Drop Down Menu ######### */<br />
<br />
.dropmenudiv_a{<br />
position:absolute;<br />
top: 0;<br />
border: 1px solid black; /*THEME CHANGE HERE*/<br />
border-top-width: 8px; /*Top border width. Should match height of .ddcolortabsline above*/<br />
border-bottom-width: 0;<br />
font:normal 10px Arial;<br />
line-height:18px;<br />
z-index:100;<br />
background-color: white;<br />
width: 200px;<br />
visibility: hidden;<br />
}<br />
<br />
<br />
.dropmenudiv_a a{<br />
width: auto;<br />
display: block;<br />
text-indent: 5px;<br />
border-top: 0 solid #678b3f;<br />
border-bottom: 1px solid #678b3f; /*THEME CHANGE HERE*/<br />
padding: 2px 0;<br />
text-decoration: none;<br />
color: black;<br />
}<br />
<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
width: 100%;<br />
}<br />
<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
background-color: #8a3c3d;<br />
color: white;<br />
}<br />
</style><br />
</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>Future Applications</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Practices</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
</ul><br />
</div><br />
<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
//SYNTAX: tabdropdown.init("menu_id", [integer OR "auto"])<br />
tabdropdown.init("colortab", 3)<br />
</script><br />
</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:11:07Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
#welcome a:visited{<br />
color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
background-position:center top;<br />
background-repeat:no-repeat;<br />
background-color:#009443;<br />
<br />
}<br />
<br />
/*Browser History*/<br />
a:link {ccolor:#20548f;}<br />
a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
a:active {color:#20548f;}<br />
<br />
#menubar.top-menu {<br />
position: absolute;<br />
top:0px;<br />
left:0px;<br />
}<br />
<br />
#menubar.slide-menu {<br />
position: absolute;<br />
top:0px;<br />
right:0px;<br />
}<br />
/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
color: #1D1D1B;<br />
}<br />
<br />
<br />
/*Code to remove/overlay standard iGEM Header for blank canvas*/<br />
#top-section{<br />
background-color: #009443;<br />
height: 100px;<br />
border: none;<br />
width: 975px;<br />
height:20px<br />
}<br />
<br />
.left-menu:hover {<br />
background-color: transparent;<br />
}<br />
<br />
.right-menu li a, .right-menu li a:hover {<br />
color: #dddddd;<br />
background-color: transparent;<br />
}<br />
<br />
<br />
<br />
#menubar {<br />
width: auto;<br />
}<br />
<br />
#p-logo{<br />
position: absolute;<br />
display: none;<br />
}<br />
<br />
#search-controls{<br />
display: none;<br />
}<br />
<br />
.firstHeading{<br />
display: none;<br />
}<br />
<br />
a{<br />
outline:none;<br />
}<br />
<br />
/*Page Controls(how wide the content section is within the window - background colour)*/ <br />
#content {<br />
width: 950x;<br />
border: none;<br />
font-family: helvetica, arial, sans-serif;<br />
color: #555555;<br />
}<br />
<br />
#footer-box {<br />
width:1075px;<br />
}<br />
</style><br />
<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
<br />
var tabdropdown={<br />
disappeardelay: 200, //set delay in miliseconds before menu disappears onmouseout<br />
disablemenuclick: false, //when user clicks on a menu item with a drop down menu, disable menu item's link?<br />
enableiframeshim: 1, //1 or 0, for true or false<br />
<br />
//No need to edit beyond here////////////////////////<br />
dropmenuobj: null, ie: document.all, firefox: document.getElementById&&!document.all, previousmenuitem:null,<br />
currentpageurl: window.location.href.replace("http://"+window.location.hostname, "").replace(/^\//, ""), //get current page url (minus hostname, ie: http://www.dynamicdrive.com/)<br />
<br />
getposOffset:function(what, offsettype){<br />
var totaloffset=(offsettype=="center")? what.offsetcenter : what.offsetTop;<br />
var parentEl=what.offsetParent;<br />
while (parentEl!=null){<br />
totaloffset=(offsettype=="center")? totaloffset+parentEl.offsetcenter : totaloffset+parentEl.offsetTop;<br />
parentEl=parentEl.offsetParent;<br />
}<br />
return totaloffset;<br />
},<br />
<br />
showhide:function(obj, e, obj2){ //obj refers to drop down menu, obj2 refers to tab menu item mouse is currently over<br />
if (this.ie || this.firefox)<br />
this.dropmenuobj.style.center=this.dropmenuobj.style.top="-500px"<br />
if (e.type=="click" && obj.visibility==hidden || e.type=="mouseover"){<br />
if (obj2.parentNode.className.indexOf("default")==-1) //if tab isn't a default selected one<br />
obj2.parentNode.className="selected"<br />
obj.visibility="visible"<br />
}<br />
else if (e.type=="click")<br />
obj.visibility="hidden"<br />
},<br />
<br />
iecompattest:function(){<br />
return (document.compatMode && document.compatMode!="BackCompat")? document.documentElement : document.body<br />
},<br />
<br />
clearbrowseredge:function(obj, whichedge){<br />
var edgeoffset=0<br />
if (whichedge=="rightedge"){<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollLeft+this.standardbody.clientWidth-15 : window.pageXOffset+window.innerWidth-15<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetWidth<br />
if (windowedge-this.dropmenuobj.x < this.dropmenuobj.contentmeasure) //move menu to the left?<br />
edgeoffset=this.dropmenuobj.contentmeasure-obj.offsetWidth<br />
}<br />
else{<br />
var topedge=this.ie && !window.opera? this.standardbody.scrollTop : window.pageYOffset<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollTop+this.standardbody.clientHeight-15 : window.pageYOffset+window.innerHeight-18<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetHeight<br />
if (windowedge-this.dropmenuobj.y < this.dropmenuobj.contentmeasure){ //move up?<br />
edgeoffset=this.dropmenuobj.contentmeasure+obj.offsetHeight<br />
if ((this.dropmenuobj.y-topedge)<this.dropmenuobj.contentmeasure) //up no good either?<br />
edgeoffset=this.dropmenuobj.y+obj.offsetHeight-topedge<br />
}<br />
this.dropmenuobj.firstlink.style.borderTopWidth=(edgeoffset==0)? 0 : "1px" //Add 1px top border to menu if dropping up<br />
}<br />
return edgeoffset<br />
},<br />
<br />
dropit:function(obj, e, dropmenuID){<br />
if (this.dropmenuobj!=null){ //hide previous menu<br />
this.dropmenuobj.style.visibility="hidden" //hide menu<br />
if (this.previousmenuitem!=null && this.previousmenuitem!=obj){<br />
if (this.previousmenuitem.parentNode.className.indexOf("default")==-1) //If the tab isn't a default selected one<br />
this.previousmenuitem.parentNode.className=""<br />
}<br />
}<br />
this.clearhidemenu()<br />
if (this.ie||this.firefox){<br />
obj.onmouseout=function(){tabdropdown.delayhidemenu(obj)}<br />
obj.onclick=function(){return !tabdropdown.disablemenuclick} //disable main menu item link onclick?<br />
this.dropmenuobj=document.getElementById(dropmenuID)<br />
this.dropmenuobj.onmouseover=function(){tabdropdown.clearhidemenu()}<br />
this.dropmenuobj.onmouseout=function(e){tabdropdown.dynamichide(e, obj)}<br />
this.dropmenuobj.onclick=function(){tabdropdown.delayhidemenu(obj)}<br />
this.showhide(this.dropmenuobj.style, e, obj)<br />
this.dropmenuobj.x=this.getposOffset(obj, "center")<br />
this.dropmenuobj.y=this.getposOffset(obj, "top")<br />
this.dropmenuobj.style.left=this.dropmenuobj.x-this.clearbrowseredge(obj, "rightedge")+"px"<br />
this.dropmenuobj.style.top=this.dropmenuobj.y-this.clearbrowseredge(obj, "bottomedge")+obj.offsetHeight+1+"px"<br />
this.previousmenuitem=obj //remember main menu item mouse moved out from (and into current menu item)<br />
this.positionshim() //call iframe shim function<br />
}<br />
},<br />
<br />
contains_firefox:function(a, b) {<br />
while (b.parentNode)<br />
if ((b = b.parentNode) == a)<br />
return true;<br />
return false;<br />
},<br />
<br />
dynamichide:function(e, obj2){ //obj2 refers to tab menu item mouse is currently over<br />
var evtobj=window.event? window.event : e<br />
if (this.ie&&!this.dropmenuobj.contains(evtobj.toElement))<br />
this.delayhidemenu(obj2)<br />
else if (this.firefox&&e.currentTarget!= evtobj.relatedTarget&& !this.contains_firefox(evtobj.currentTarget, evtobj.relatedTarget))<br />
this.delayhidemenu(obj2)<br />
},<br />
<br />
delayhidemenu:function(obj2){<br />
this.delayhide=setTimeout(function(){tabdropdown.dropmenuobj.style.visibility='hidden'; if (obj2.parentNode.className.indexOf('default')==-1) obj2.parentNode.className=''},this.disappeardelay) //hide menu<br />
},<br />
<br />
clearhidemenu:function(){<br />
if (this.delayhide!="undefined")<br />
clearTimeout(this.delayhide)<br />
},<br />
<br />
positionshim:function(){ //display iframe shim function<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined"){<br />
if (this.dropmenuobj.style.visibility=="visible"){<br />
this.shimobject.style.width=this.dropmenuobj.offsetWidth+"px"<br />
this.shimobject.style.height=this.dropmenuobj.offsetHeight+"px"<br />
this.shimobject.style.center=this.dropmenuobj.style.center<br />
this.shimobject.style.top=this.dropmenuobj.style.top<br />
}<br />
this.shimobject.style.display=(this.dropmenuobj.style.visibility=="visible")? "block" : "none"<br />
}<br />
},<br />
<br />
hideshim:function(){<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined")<br />
this.shimobject.style.display='none'<br />
},<br />
<br />
isSelected:function(menuurl){<br />
var menuurl=menuurl.replace("http://"+menuurl.hostname, "").replace(/^\//, "")<br />
return (tabdropdown.currentpageurl==menuurl)<br />
},<br />
<br />
init:function(menuid, dselected){<br />
this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
var menuitems=document.getElementById(menuid).getElementsByTagName("a")<br />
for (var i=0; i<menuitems.length; i++){<br />
if (menuitems[i].getAttribute("rel")){<br />
var relvalue=menuitems[i].getAttribute("rel")<br />
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/* ######### Style for Drop Down Menu ######### */<br />
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<br />
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<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
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<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
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</style><br />
</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>Future Applications</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Practices</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
</ul><br />
</div><br />
<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
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</script><br />
</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:10:31Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
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color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
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<br />
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/*Browser History*/<br />
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#menubar.slide-menu {<br />
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/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
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<br />
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<br />
<br />
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a{<br />
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<br />
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<br />
#footer-box {<br />
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<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
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<br />
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this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
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<br />
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<br />
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<br />
.ddcolortabs a:hover span{<br />
background-color: #00632d;<br />
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<br />
.ddcolortabs .selected a, #ddcolortabs .selected a span{ /*currently selected tab*/<br />
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<br />
.ddcolortabsline{<br />
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padding: 0;<br />
width: 100%;<br />
height: 8px;<br />
line-height: 8px;<br />
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}<br />
<br />
/* ######### Style for Drop Down Menu ######### */<br />
<br />
.dropmenudiv_a{<br />
position:absolute;<br />
top: 0;<br />
border: 1px solid black; /*THEME CHANGE HERE*/<br />
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border-bottom-width: 0;<br />
font:normal 10px Arial;<br />
line-height:18px;<br />
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<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
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<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
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color: white;<br />
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</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>Future Applications</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
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<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
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</script><br />
</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:09:50Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
#welcome a:visited{<br />
color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
background-position:center top;<br />
background-repeat:no-repeat;<br />
background-color:#009443;<br />
<br />
}<br />
<br />
/*Browser History*/<br />
a:link {ccolor:#20548f;}<br />
a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
a:active {color:#20548f;}<br />
<br />
#menubar.top-menu {<br />
position: absolute;<br />
top:0px;<br />
left:0px;<br />
}<br />
<br />
#menubar.slide-menu {<br />
position: absolute;<br />
top:0px;<br />
right:0px;<br />
}<br />
/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
color: #1D1D1B;<br />
}<br />
<br />
<br />
/*Code to remove/overlay standard iGEM Header for blank canvas*/<br />
#top-section{<br />
background-color: #009443;<br />
height: 100px;<br />
border: none;<br />
width: 975px;<br />
height:20px<br />
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.left-menu:hover {<br />
background-color: transparent;<br />
}<br />
<br />
.right-menu li a, .right-menu li a:hover {<br />
color: #dddddd;<br />
background-color: transparent;<br />
}<br />
<br />
<br />
<br />
#menubar {<br />
width: auto;<br />
}<br />
<br />
#p-logo{<br />
position: absolute;<br />
display: none;<br />
}<br />
<br />
#search-controls{<br />
display: none;<br />
}<br />
<br />
.firstHeading{<br />
display: none;<br />
}<br />
<br />
a{<br />
outline:none;<br />
}<br />
<br />
/*Page Controls(how wide the content section is within the window - background colour)*/ <br />
#content {<br />
width: 950x;<br />
border: none;<br />
font-family: helvetica, arial, sans-serif;<br />
color: #555555;<br />
}<br />
<br />
#footer-box {<br />
width:1075px;<br />
}<br />
</style><br />
<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
<br />
var tabdropdown={<br />
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enableiframeshim: 1, //1 or 0, for true or false<br />
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//No need to edit beyond here////////////////////////<br />
dropmenuobj: null, ie: document.all, firefox: document.getElementById&&!document.all, previousmenuitem:null,<br />
currentpageurl: window.location.href.replace("http://"+window.location.hostname, "").replace(/^\//, ""), //get current page url (minus hostname, ie: http://www.dynamicdrive.com/)<br />
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<br />
showhide:function(obj, e, obj2){ //obj refers to drop down menu, obj2 refers to tab menu item mouse is currently over<br />
if (this.ie || this.firefox)<br />
this.dropmenuobj.style.center=this.dropmenuobj.style.top="-500px"<br />
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obj.visibility="hidden"<br />
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dropit:function(obj, e, dropmenuID){<br />
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this.clearhidemenu()<br />
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obj.onmouseout=function(){tabdropdown.delayhidemenu(obj)}<br />
obj.onclick=function(){return !tabdropdown.disablemenuclick} //disable main menu item link onclick?<br />
this.dropmenuobj=document.getElementById(dropmenuID)<br />
this.dropmenuobj.onmouseover=function(){tabdropdown.clearhidemenu()}<br />
this.dropmenuobj.onmouseout=function(e){tabdropdown.dynamichide(e, obj)}<br />
this.dropmenuobj.onclick=function(){tabdropdown.delayhidemenu(obj)}<br />
this.showhide(this.dropmenuobj.style, e, obj)<br />
this.dropmenuobj.x=this.getposOffset(obj, "center")<br />
this.dropmenuobj.y=this.getposOffset(obj, "top")<br />
this.dropmenuobj.style.left=this.dropmenuobj.x-this.clearbrowseredge(obj, "rightedge")+"px"<br />
this.dropmenuobj.style.top=this.dropmenuobj.y-this.clearbrowseredge(obj, "bottomedge")+obj.offsetHeight+1+"px"<br />
this.previousmenuitem=obj //remember main menu item mouse moved out from (and into current menu item)<br />
this.positionshim() //call iframe shim function<br />
}<br />
},<br />
<br />
contains_firefox:function(a, b) {<br />
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if ((b = b.parentNode) == a)<br />
return true;<br />
return false;<br />
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<br />
dynamichide:function(e, obj2){ //obj2 refers to tab menu item mouse is currently over<br />
var evtobj=window.event? window.event : e<br />
if (this.ie&&!this.dropmenuobj.contains(evtobj.toElement))<br />
this.delayhidemenu(obj2)<br />
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this.delayhidemenu(obj2)<br />
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<br />
delayhidemenu:function(obj2){<br />
this.delayhide=setTimeout(function(){tabdropdown.dropmenuobj.style.visibility='hidden'; if (obj2.parentNode.className.indexOf('default')==-1) obj2.parentNode.className=''},this.disappeardelay) //hide menu<br />
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hideshim:function(){<br />
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<br />
isSelected:function(menuurl){<br />
var menuurl=menuurl.replace("http://"+menuurl.hostname, "").replace(/^\//, "")<br />
return (tabdropdown.currentpageurl==menuurl)<br />
},<br />
<br />
init:function(menuid, dselected){<br />
this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
var menuitems=document.getElementById(menuid).getElementsByTagName("a")<br />
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if (dselected=="auto" && typeof setalready=="undefined" && this.isSelected(menuitems[i].href)){<br />
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menuitems[i].parentNode.className+=" selected default"<br />
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<br />
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<style><br />
.ddcolortabs{<br />
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<br />
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color: white;<br />
background: #009443 url(media/color_tabs_left.gif) no-repeat left top;<br />
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text-decoration:none;<br />
letter-spacing: 1px;<br />
}<br />
<br />
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display:block;<br />
background: transparent url(media/color_tabs_right.gif) no-repeat right top;<br />
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<br />
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}<br />
<br />
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<br />
.ddcolortabs a:hover span{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs .selected a, #ddcolortabs .selected a span{ /*currently selected tab*/<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabsline{<br />
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padding: 0;<br />
width: 100%;<br />
height: 8px;<br />
line-height: 8px;<br />
background: #009443;<br />
border-top: 1px solid #fff; /*Remove this to remove border between bar and tabs*/<br />
}<br />
<br />
/* ######### Style for Drop Down Menu ######### */<br />
<br />
.dropmenudiv_a{<br />
position:absolute;<br />
top: 0;<br />
border: 1px solid black; /*THEME CHANGE HERE*/<br />
border-top-width: 8px; /*Top border width. Should match height of .ddcolortabsline above*/<br />
border-bottom-width: 0;<br />
font:normal 10px Arial;<br />
line-height:18px;<br />
z-index:100;<br />
background-color: white;<br />
width: 200px;<br />
visibility: hidden;<br />
}<br />
<br />
<br />
.dropmenudiv_a a{<br />
width: auto;<br />
display: block;<br />
text-indent: 5px;<br />
border-top: 0 solid #678b3f;<br />
border-bottom: 1px solid #678b3f; /*THEME CHANGE HERE*/<br />
padding: 2px 0;<br />
text-decoration: none;<br />
color: black;<br />
}<br />
<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
width: 100%;<br />
}<br />
<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
background-color: #8a3c3d;<br />
color: white;<br />
}<br />
</style><br />
</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>The Future</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Practices</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
</ul><br />
</div><br />
<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
//SYNTAX: tabdropdown.init("menu_id", [integer OR "auto"])<br />
tabdropdown.init("colortab", 3)<br />
</script><br />
</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:09:13Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
#welcome a:visited{<br />
color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
background-position:center top;<br />
background-repeat:no-repeat;<br />
background-color:#009443;<br />
<br />
}<br />
<br />
/*Browser History*/<br />
a:link {ccolor:#20548f;}<br />
a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
a:active {color:#20548f;}<br />
<br />
#menubar.top-menu {<br />
position: absolute;<br />
top:0px;<br />
left:0px;<br />
}<br />
<br />
#menubar.slide-menu {<br />
position: absolute;<br />
top:0px;<br />
right:0px;<br />
}<br />
/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
color: #1D1D1B;<br />
}<br />
<br />
<br />
/*Code to remove/overlay standard iGEM Header for blank canvas*/<br />
#top-section{<br />
background-color: #009443;<br />
height: 100px;<br />
border: none;<br />
width: 975px;<br />
height:20px<br />
}<br />
<br />
.left-menu:hover {<br />
background-color: transparent;<br />
}<br />
<br />
.right-menu li a, .right-menu li a:hover {<br />
color: #dddddd;<br />
background-color: transparent;<br />
}<br />
<br />
<br />
<br />
#menubar {<br />
width: auto;<br />
}<br />
<br />
#p-logo{<br />
position: absolute;<br />
display: none;<br />
}<br />
<br />
#search-controls{<br />
display: none;<br />
}<br />
<br />
.firstHeading{<br />
display: none;<br />
}<br />
<br />
a{<br />
outline:none;<br />
}<br />
<br />
/*Page Controls(how wide the content section is within the window - background colour)*/ <br />
#content {<br />
width: 950x;<br />
border: none;<br />
font-family: helvetica, arial, sans-serif;<br />
color: #555555;<br />
}<br />
<br />
#footer-box {<br />
width:1075px;<br />
}<br />
</style><br />
<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
<br />
var tabdropdown={<br />
disappeardelay: 200, //set delay in miliseconds before menu disappears onmouseout<br />
disablemenuclick: false, //when user clicks on a menu item with a drop down menu, disable menu item's link?<br />
enableiframeshim: 1, //1 or 0, for true or false<br />
<br />
//No need to edit beyond here////////////////////////<br />
dropmenuobj: null, ie: document.all, firefox: document.getElementById&&!document.all, previousmenuitem:null,<br />
currentpageurl: window.location.href.replace("http://"+window.location.hostname, "").replace(/^\//, ""), //get current page url (minus hostname, ie: http://www.dynamicdrive.com/)<br />
<br />
getposOffset:function(what, offsettype){<br />
var totaloffset=(offsettype=="center")? what.offsetcenter : what.offsetTop;<br />
var parentEl=what.offsetParent;<br />
while (parentEl!=null){<br />
totaloffset=(offsettype=="center")? totaloffset+parentEl.offsetcenter : totaloffset+parentEl.offsetTop;<br />
parentEl=parentEl.offsetParent;<br />
}<br />
return totaloffset;<br />
},<br />
<br />
showhide:function(obj, e, obj2){ //obj refers to drop down menu, obj2 refers to tab menu item mouse is currently over<br />
if (this.ie || this.firefox)<br />
this.dropmenuobj.style.center=this.dropmenuobj.style.top="-500px"<br />
if (e.type=="click" && obj.visibility==hidden || e.type=="mouseover"){<br />
if (obj2.parentNode.className.indexOf("default")==-1) //if tab isn't a default selected one<br />
obj2.parentNode.className="selected"<br />
obj.visibility="visible"<br />
}<br />
else if (e.type=="click")<br />
obj.visibility="hidden"<br />
},<br />
<br />
iecompattest:function(){<br />
return (document.compatMode && document.compatMode!="BackCompat")? document.documentElement : document.body<br />
},<br />
<br />
clearbrowseredge:function(obj, whichedge){<br />
var edgeoffset=0<br />
if (whichedge=="rightedge"){<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollLeft+this.standardbody.clientWidth-15 : window.pageXOffset+window.innerWidth-15<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetWidth<br />
if (windowedge-this.dropmenuobj.x < this.dropmenuobj.contentmeasure) //move menu to the left?<br />
edgeoffset=this.dropmenuobj.contentmeasure-obj.offsetWidth<br />
}<br />
else{<br />
var topedge=this.ie && !window.opera? this.standardbody.scrollTop : window.pageYOffset<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollTop+this.standardbody.clientHeight-15 : window.pageYOffset+window.innerHeight-18<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetHeight<br />
if (windowedge-this.dropmenuobj.y < this.dropmenuobj.contentmeasure){ //move up?<br />
edgeoffset=this.dropmenuobj.contentmeasure+obj.offsetHeight<br />
if ((this.dropmenuobj.y-topedge)<this.dropmenuobj.contentmeasure) //up no good either?<br />
edgeoffset=this.dropmenuobj.y+obj.offsetHeight-topedge<br />
}<br />
this.dropmenuobj.firstlink.style.borderTopWidth=(edgeoffset==0)? 0 : "1px" //Add 1px top border to menu if dropping up<br />
}<br />
return edgeoffset<br />
},<br />
<br />
dropit:function(obj, e, dropmenuID){<br />
if (this.dropmenuobj!=null){ //hide previous menu<br />
this.dropmenuobj.style.visibility="hidden" //hide menu<br />
if (this.previousmenuitem!=null && this.previousmenuitem!=obj){<br />
if (this.previousmenuitem.parentNode.className.indexOf("default")==-1) //If the tab isn't a default selected one<br />
this.previousmenuitem.parentNode.className=""<br />
}<br />
}<br />
this.clearhidemenu()<br />
if (this.ie||this.firefox){<br />
obj.onmouseout=function(){tabdropdown.delayhidemenu(obj)}<br />
obj.onclick=function(){return !tabdropdown.disablemenuclick} //disable main menu item link onclick?<br />
this.dropmenuobj=document.getElementById(dropmenuID)<br />
this.dropmenuobj.onmouseover=function(){tabdropdown.clearhidemenu()}<br />
this.dropmenuobj.onmouseout=function(e){tabdropdown.dynamichide(e, obj)}<br />
this.dropmenuobj.onclick=function(){tabdropdown.delayhidemenu(obj)}<br />
this.showhide(this.dropmenuobj.style, e, obj)<br />
this.dropmenuobj.x=this.getposOffset(obj, "center")<br />
this.dropmenuobj.y=this.getposOffset(obj, "top")<br />
this.dropmenuobj.style.left=this.dropmenuobj.x-this.clearbrowseredge(obj, "rightedge")+"px"<br />
this.dropmenuobj.style.top=this.dropmenuobj.y-this.clearbrowseredge(obj, "bottomedge")+obj.offsetHeight+1+"px"<br />
this.previousmenuitem=obj //remember main menu item mouse moved out from (and into current menu item)<br />
this.positionshim() //call iframe shim function<br />
}<br />
},<br />
<br />
contains_firefox:function(a, b) {<br />
while (b.parentNode)<br />
if ((b = b.parentNode) == a)<br />
return true;<br />
return false;<br />
},<br />
<br />
dynamichide:function(e, obj2){ //obj2 refers to tab menu item mouse is currently over<br />
var evtobj=window.event? window.event : e<br />
if (this.ie&&!this.dropmenuobj.contains(evtobj.toElement))<br />
this.delayhidemenu(obj2)<br />
else if (this.firefox&&e.currentTarget!= evtobj.relatedTarget&& !this.contains_firefox(evtobj.currentTarget, evtobj.relatedTarget))<br />
this.delayhidemenu(obj2)<br />
},<br />
<br />
delayhidemenu:function(obj2){<br />
this.delayhide=setTimeout(function(){tabdropdown.dropmenuobj.style.visibility='hidden'; if (obj2.parentNode.className.indexOf('default')==-1) obj2.parentNode.className=''},this.disappeardelay) //hide menu<br />
},<br />
<br />
clearhidemenu:function(){<br />
if (this.delayhide!="undefined")<br />
clearTimeout(this.delayhide)<br />
},<br />
<br />
positionshim:function(){ //display iframe shim function<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined"){<br />
if (this.dropmenuobj.style.visibility=="visible"){<br />
this.shimobject.style.width=this.dropmenuobj.offsetWidth+"px"<br />
this.shimobject.style.height=this.dropmenuobj.offsetHeight+"px"<br />
this.shimobject.style.center=this.dropmenuobj.style.center<br />
this.shimobject.style.top=this.dropmenuobj.style.top<br />
}<br />
this.shimobject.style.display=(this.dropmenuobj.style.visibility=="visible")? "block" : "none"<br />
}<br />
},<br />
<br />
hideshim:function(){<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined")<br />
this.shimobject.style.display='none'<br />
},<br />
<br />
isSelected:function(menuurl){<br />
var menuurl=menuurl.replace("http://"+menuurl.hostname, "").replace(/^\//, "")<br />
return (tabdropdown.currentpageurl==menuurl)<br />
},<br />
<br />
init:function(menuid, dselected){<br />
this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
var menuitems=document.getElementById(menuid).getElementsByTagName("a")<br />
for (var i=0; i<menuitems.length; i++){<br />
if (menuitems[i].getAttribute("rel")){<br />
var relvalue=menuitems[i].getAttribute("rel")<br />
document.getElementById(relvalue).firstlink=document.getElementById(relvalue).getElementsByTagName("a")[0]<br />
menuitems[i].onmouseover=function(e){<br />
var event=typeof e!="undefined"? e : window.event<br />
tabdropdown.dropit(this, event, this.getAttribute("rel"))<br />
}<br />
}<br />
if (dselected=="auto" && typeof setalready=="undefined" && this.isSelected(menuitems[i].href)){<br />
menuitems[i].parentNode.className+=" selected default"<br />
var setalready=true<br />
}<br />
else if (parseInt(dselected)==i)<br />
menuitems[i].parentNode.className+=" selected default"<br />
}<br />
}<br />
<br />
}<br />
</script><br />
<style><br />
.ddcolortabs{<br />
padding: 0;<br />
width: 100%;<br />
background: transparent;<br />
voice-family: "\"}\"";<br />
voice-family: inherit;<br />
}<br />
<br />
.ddcolortabs ul{<br />
font: normal 16px Calibri, sans-serif;<br />
margin:0;<br />
padding:0;<br />
list-style:none;<br />
}<br />
<br />
.ddcolortabs li{<br />
display:inline;<br />
margin:0 2px 0 0;<br />
padding:0;<br />
text-transform:uppercase;<br />
}<br />
<br />
<br />
.ddcolortabs a{<br />
float: left;<br />
color: white;<br />
background: #009443 url(media/color_tabs_left.gif) no-repeat left top;<br />
margin:0 2px 0 0;<br />
padding:0 0 1px 2px;<br />
text-decoration:none;<br />
letter-spacing: 1px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:left;<br />
display:block;<br />
background: transparent url(media/color_tabs_right.gif) no-repeat right top;<br />
padding: 4px 8px 2px 7px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:none;<br />
}<br />
<br />
.ddcolortabs a:hover{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs a:hover span{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs .selected a, #ddcolortabs .selected a span{ /*currently selected tab*/<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabsline{<br />
clear: both;<br />
padding: 0;<br />
width: 100%;<br />
height: 8px;<br />
line-height: 8px;<br />
background: #009443;<br />
border-top: 1px solid #fff; /*Remove this to remove border between bar and tabs*/<br />
}<br />
<br />
/* ######### Style for Drop Down Menu ######### */<br />
<br />
.dropmenudiv_a{<br />
position:absolute;<br />
top: 0;<br />
border: 1px solid black; /*THEME CHANGE HERE*/<br />
border-top-width: 8px; /*Top border width. Should match height of .ddcolortabsline above*/<br />
border-bottom-width: 0;<br />
font:normal 12px Arial;<br />
line-height:18px;<br />
z-index:100;<br />
background-color: white;<br />
width: 200px;<br />
visibility: hidden;<br />
}<br />
<br />
<br />
.dropmenudiv_a a{<br />
width: auto;<br />
display: block;<br />
text-indent: 5px;<br />
border-top: 0 solid #678b3f;<br />
border-bottom: 1px solid #678b3f; /*THEME CHANGE HERE*/<br />
padding: 2px 0;<br />
text-decoration: none;<br />
color: black;<br />
}<br />
<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
width: 100%;<br />
}<br />
<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
background-color: #8a3c3d;<br />
color: white;<br />
}<br />
</style><br />
</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>The Future</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Outreach</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
</ul><br />
</div><br />
<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
//SYNTAX: tabdropdown.init("menu_id", [integer OR "auto"])<br />
tabdropdown.init("colortab", 3)<br />
</script><br />
</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/Template:UEANRPTemplate:UEANRP2012-09-27T00:04:55Z<p>RussellGritton: </p>
<hr />
<div>[[Image:Header1NewGreen.png|top]]<br />
<br />
<html><style><br />
#content a[href ^="https://"].external<br />
{<br />
background: center right no-repeat; padding-right: 18px;<br />
}</style></html><br />
<br />
<html><style type="text/css"><br />
#edi-twitter-widget .plainlinks{<br />
border: 1px solid #ccc;<br />
}<br />
#welcome a,<br />
#welcome a:hover,<br />
#welcome a:visited{<br />
color: #009443;<br />
}<br />
</style></html><br />
<br />
<html><br />
<br />
<style><br />
body{<br />
background-image:url("https://static.igem.org/mediawiki/2012/5/5a/WikiBGNewGreen.jpg");<br />
background-position:center top;<br />
background-repeat:no-repeat;<br />
background-color:#009443;<br />
<br />
}<br />
<br />
/*Browser History*/<br />
a:link {ccolor:#20548f;}<br />
a:visited {color:#20548f;}<br />
a:hover {color:#20548f;}<br />
a:active {color:#20548f;}<br />
<br />
#menubar.top-menu {<br />
position: absolute;<br />
top:0px;<br />
left:0px;<br />
}<br />
<br />
#menubar.slide-menu {<br />
position: absolute;<br />
top:0px;<br />
right:0px;<br />
}<br />
/*Top Control Bar*/ <br />
#menubar a:active,#menubar a:hover,#menubar a:link,#menubar a:visited {<br />
color: #1D1D1B;<br />
}<br />
<br />
<br />
/*Code to remove/overlay standard iGEM Header for blank canvas*/<br />
#top-section{<br />
background-color: #009443;<br />
height: 100px;<br />
border: none;<br />
width: 975px;<br />
height:20px<br />
}<br />
<br />
.left-menu:hover {<br />
background-color: transparent;<br />
}<br />
<br />
.right-menu li a, .right-menu li a:hover {<br />
color: #dddddd;<br />
background-color: transparent;<br />
}<br />
<br />
<br />
<br />
#menubar {<br />
width: auto;<br />
}<br />
<br />
#p-logo{<br />
position: absolute;<br />
display: none;<br />
}<br />
<br />
#search-controls{<br />
display: none;<br />
}<br />
<br />
.firstHeading{<br />
display: none;<br />
}<br />
<br />
a{<br />
outline:none;<br />
}<br />
<br />
/*Page Controls(how wide the content section is within the window - background colour)*/ <br />
#content {<br />
width: 950x;<br />
border: none;<br />
font-family: helvetica, arial, sans-serif;<br />
color: #555555;<br />
}<br />
<br />
#footer-box {<br />
width:1075px;<br />
}<br />
</style><br />
<br />
<head><br />
<title>NRP UEA iGEM 2012</title><br />
<script type="text/javascript"><br />
<br />
var tabdropdown={<br />
disappeardelay: 200, //set delay in miliseconds before menu disappears onmouseout<br />
disablemenuclick: false, //when user clicks on a menu item with a drop down menu, disable menu item's link?<br />
enableiframeshim: 1, //1 or 0, for true or false<br />
<br />
//No need to edit beyond here////////////////////////<br />
dropmenuobj: null, ie: document.all, firefox: document.getElementById&&!document.all, previousmenuitem:null,<br />
currentpageurl: window.location.href.replace("http://"+window.location.hostname, "").replace(/^\//, ""), //get current page url (minus hostname, ie: http://www.dynamicdrive.com/)<br />
<br />
getposOffset:function(what, offsettype){<br />
var totaloffset=(offsettype=="center")? what.offsetcenter : what.offsetTop;<br />
var parentEl=what.offsetParent;<br />
while (parentEl!=null){<br />
totaloffset=(offsettype=="center")? totaloffset+parentEl.offsetcenter : totaloffset+parentEl.offsetTop;<br />
parentEl=parentEl.offsetParent;<br />
}<br />
return totaloffset;<br />
},<br />
<br />
showhide:function(obj, e, obj2){ //obj refers to drop down menu, obj2 refers to tab menu item mouse is currently over<br />
if (this.ie || this.firefox)<br />
this.dropmenuobj.style.center=this.dropmenuobj.style.top="-500px"<br />
if (e.type=="click" && obj.visibility==hidden || e.type=="mouseover"){<br />
if (obj2.parentNode.className.indexOf("default")==-1) //if tab isn't a default selected one<br />
obj2.parentNode.className="selected"<br />
obj.visibility="visible"<br />
}<br />
else if (e.type=="click")<br />
obj.visibility="hidden"<br />
},<br />
<br />
iecompattest:function(){<br />
return (document.compatMode && document.compatMode!="BackCompat")? document.documentElement : document.body<br />
},<br />
<br />
clearbrowseredge:function(obj, whichedge){<br />
var edgeoffset=0<br />
if (whichedge=="rightedge"){<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollLeft+this.standardbody.clientWidth-15 : window.pageXOffset+window.innerWidth-15<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetWidth<br />
if (windowedge-this.dropmenuobj.x < this.dropmenuobj.contentmeasure) //move menu to the left?<br />
edgeoffset=this.dropmenuobj.contentmeasure-obj.offsetWidth<br />
}<br />
else{<br />
var topedge=this.ie && !window.opera? this.standardbody.scrollTop : window.pageYOffset<br />
var windowedge=this.ie && !window.opera? this.standardbody.scrollTop+this.standardbody.clientHeight-15 : window.pageYOffset+window.innerHeight-18<br />
this.dropmenuobj.contentmeasure=this.dropmenuobj.offsetHeight<br />
if (windowedge-this.dropmenuobj.y < this.dropmenuobj.contentmeasure){ //move up?<br />
edgeoffset=this.dropmenuobj.contentmeasure+obj.offsetHeight<br />
if ((this.dropmenuobj.y-topedge)<this.dropmenuobj.contentmeasure) //up no good either?<br />
edgeoffset=this.dropmenuobj.y+obj.offsetHeight-topedge<br />
}<br />
this.dropmenuobj.firstlink.style.borderTopWidth=(edgeoffset==0)? 0 : "1px" //Add 1px top border to menu if dropping up<br />
}<br />
return edgeoffset<br />
},<br />
<br />
dropit:function(obj, e, dropmenuID){<br />
if (this.dropmenuobj!=null){ //hide previous menu<br />
this.dropmenuobj.style.visibility="hidden" //hide menu<br />
if (this.previousmenuitem!=null && this.previousmenuitem!=obj){<br />
if (this.previousmenuitem.parentNode.className.indexOf("default")==-1) //If the tab isn't a default selected one<br />
this.previousmenuitem.parentNode.className=""<br />
}<br />
}<br />
this.clearhidemenu()<br />
if (this.ie||this.firefox){<br />
obj.onmouseout=function(){tabdropdown.delayhidemenu(obj)}<br />
obj.onclick=function(){return !tabdropdown.disablemenuclick} //disable main menu item link onclick?<br />
this.dropmenuobj=document.getElementById(dropmenuID)<br />
this.dropmenuobj.onmouseover=function(){tabdropdown.clearhidemenu()}<br />
this.dropmenuobj.onmouseout=function(e){tabdropdown.dynamichide(e, obj)}<br />
this.dropmenuobj.onclick=function(){tabdropdown.delayhidemenu(obj)}<br />
this.showhide(this.dropmenuobj.style, e, obj)<br />
this.dropmenuobj.x=this.getposOffset(obj, "center")<br />
this.dropmenuobj.y=this.getposOffset(obj, "top")<br />
this.dropmenuobj.style.left=this.dropmenuobj.x-this.clearbrowseredge(obj, "rightedge")+"px"<br />
this.dropmenuobj.style.top=this.dropmenuobj.y-this.clearbrowseredge(obj, "bottomedge")+obj.offsetHeight+1+"px"<br />
this.previousmenuitem=obj //remember main menu item mouse moved out from (and into current menu item)<br />
this.positionshim() //call iframe shim function<br />
}<br />
},<br />
<br />
contains_firefox:function(a, b) {<br />
while (b.parentNode)<br />
if ((b = b.parentNode) == a)<br />
return true;<br />
return false;<br />
},<br />
<br />
dynamichide:function(e, obj2){ //obj2 refers to tab menu item mouse is currently over<br />
var evtobj=window.event? window.event : e<br />
if (this.ie&&!this.dropmenuobj.contains(evtobj.toElement))<br />
this.delayhidemenu(obj2)<br />
else if (this.firefox&&e.currentTarget!= evtobj.relatedTarget&& !this.contains_firefox(evtobj.currentTarget, evtobj.relatedTarget))<br />
this.delayhidemenu(obj2)<br />
},<br />
<br />
delayhidemenu:function(obj2){<br />
this.delayhide=setTimeout(function(){tabdropdown.dropmenuobj.style.visibility='hidden'; if (obj2.parentNode.className.indexOf('default')==-1) obj2.parentNode.className=''},this.disappeardelay) //hide menu<br />
},<br />
<br />
clearhidemenu:function(){<br />
if (this.delayhide!="undefined")<br />
clearTimeout(this.delayhide)<br />
},<br />
<br />
positionshim:function(){ //display iframe shim function<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined"){<br />
if (this.dropmenuobj.style.visibility=="visible"){<br />
this.shimobject.style.width=this.dropmenuobj.offsetWidth+"px"<br />
this.shimobject.style.height=this.dropmenuobj.offsetHeight+"px"<br />
this.shimobject.style.center=this.dropmenuobj.style.center<br />
this.shimobject.style.top=this.dropmenuobj.style.top<br />
}<br />
this.shimobject.style.display=(this.dropmenuobj.style.visibility=="visible")? "block" : "none"<br />
}<br />
},<br />
<br />
hideshim:function(){<br />
if (this.enableiframeshim && typeof this.shimobject!="undefined")<br />
this.shimobject.style.display='none'<br />
},<br />
<br />
isSelected:function(menuurl){<br />
var menuurl=menuurl.replace("http://"+menuurl.hostname, "").replace(/^\//, "")<br />
return (tabdropdown.currentpageurl==menuurl)<br />
},<br />
<br />
init:function(menuid, dselected){<br />
this.standardbody=(document.compatMode=="CSS1Compat")? document.documentElement : document.body //create reference to common "body" across doctypes<br />
var menuitems=document.getElementById(menuid).getElementsByTagName("a")<br />
for (var i=0; i<menuitems.length; i++){<br />
if (menuitems[i].getAttribute("rel")){<br />
var relvalue=menuitems[i].getAttribute("rel")<br />
document.getElementById(relvalue).firstlink=document.getElementById(relvalue).getElementsByTagName("a")[0]<br />
menuitems[i].onmouseover=function(e){<br />
var event=typeof e!="undefined"? e : window.event<br />
tabdropdown.dropit(this, event, this.getAttribute("rel"))<br />
}<br />
}<br />
if (dselected=="auto" && typeof setalready=="undefined" && this.isSelected(menuitems[i].href)){<br />
menuitems[i].parentNode.className+=" selected default"<br />
var setalready=true<br />
}<br />
else if (parseInt(dselected)==i)<br />
menuitems[i].parentNode.className+=" selected default"<br />
}<br />
}<br />
<br />
}<br />
</script><br />
<style><br />
.ddcolortabs{<br />
padding: 0;<br />
width: 100%;<br />
background: transparent;<br />
voice-family: "\"}\"";<br />
voice-family: inherit;<br />
}<br />
<br />
.ddcolortabs ul{<br />
font: normal 16px Calibri, sans-serif;<br />
margin:0;<br />
padding:0;<br />
list-style:none;<br />
}<br />
<br />
.ddcolortabs li{<br />
display:inline;<br />
margin:0 2px 0 0;<br />
padding:0;<br />
text-transform:uppercase;<br />
}<br />
<br />
<br />
.ddcolortabs a{<br />
float: left;<br />
color: white;<br />
background: #009443 url(media/color_tabs_left.gif) no-repeat left top;<br />
margin:0 2px 0 0;<br />
padding:0 0 1px 2px;<br />
text-decoration:none;<br />
letter-spacing: 1px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:left;<br />
display:block;<br />
background: transparent url(media/color_tabs_right.gif) no-repeat right top;<br />
padding: 4px 8px 2px 7px;<br />
}<br />
<br />
.ddcolortabs a span{<br />
float:none;<br />
}<br />
<br />
.ddcolortabs a:hover{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs a:hover span{<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabs .selected a, #ddcolortabs .selected a span{ /*currently selected tab*/<br />
background-color: #00632d;<br />
}<br />
<br />
.ddcolortabsline{<br />
clear: both;<br />
padding: 0;<br />
width: 100%;<br />
height: 8px;<br />
line-height: 8px;<br />
background: #009443;<br />
border-top: 1px solid #fff; /*Remove this to remove border between bar and tabs*/<br />
}<br />
<br />
/* ######### Style for Drop Down Menu ######### */<br />
<br />
.dropmenudiv_a{<br />
position:absolute;<br />
top: 0;<br />
border: 1px solid black; /*THEME CHANGE HERE*/<br />
border-top-width: 8px; /*Top border width. Should match height of .ddcolortabsline above*/<br />
border-bottom-width: 0;<br />
font:normal 12px Arial;<br />
line-height:18px;<br />
z-index:100;<br />
background-color: white;<br />
width: 200px;<br />
visibility: hidden;<br />
}<br />
<br />
<br />
.dropmenudiv_a a{<br />
width: auto;<br />
display: block;<br />
text-indent: 5px;<br />
border-top: 0 solid #678b3f;<br />
border-bottom: 1px solid #678b3f; /*THEME CHANGE HERE*/<br />
padding: 2px 0;<br />
text-decoration: none;<br />
color: black;<br />
}<br />
<br />
* html .dropmenudiv_a a{ /*IE only hack*/<br />
width: 100%;<br />
}<br />
<br />
.dropmenudiv_a a:hover{ /*THEME CHANGE HERE*/<br />
background-color: #8a3c3d;<br />
color: white;<br />
}<br />
</style><br />
</head><br />
<!-- CSS for Drop Down Tabs Menu #1 --><br />
<link rel="stylesheet" type="text/css" href="ddcolortabs.css" /><br />
<br />
<div id="colortab" class="ddcolortabs"><br />
<ul><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich" title="Home"><span>Home</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Team" rel="dropmenu1_a"><span>Team</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Project" title="Home"><span>Project</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Safety" rel="dropmenu1_a"><span>Safety</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Notebook" rel="dropmenu1_a"><span>Lab Book</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Parts" rel="dropmenu1_a"><span>Parts</span></a></li><br />
<br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Future" rel="dropmenu1_a"><span>The Future</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare" rel="dropmenu1_a"><span>Quanticare</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Human Outreach" rel="dropmenu1_a"><span>Human Practices</span></a></li><br />
<li><a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Attributions" rel="dropmenu1_a"><span>Attributions</span></a></li><br />
</ul><br />
</div><br />
<div class="ddcolortabsline">&nbsp;</div><br />
<div id="dropmenu1_a" class="dropmenudiv_a"><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Overview</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step1</a><br />
<a href="https://2008.igem.org/Team:Edinburgh/Team">Step2</a><br />
</div><br />
<script type="text/javascript"><br />
//SYNTAX: tabdropdown.init("menu_id", [integer OR "auto"])<br />
tabdropdown.init("colortab", 3)<br />
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</body><br />
</html></div>RussellGrittonhttp://2012.igem.org/File:WikiBGNewGreen.jpgFile:WikiBGNewGreen.jpg2012-09-27T00:03:59Z<p>RussellGritton: </p>
<hr />
<div></div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/ProjectTeam:NRP-UEA-Norwich/Project2012-09-26T23:46:06Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRP}}<br />
<br />
==NRP-UEA-Norwich Project Overview==<br />
<br />
Sensory BioBrick systems have been a large constituent of previous iGEM projects in which teams have combined impressive amounts of logic with limitless creativity in order to produce synthetically engineered organisms with the ability to detect the presence of specific substrates; this was achieved by combining various promoters and reporters to produce novel gene systems of great breadth and depth.<br />
<br />
We too have taken a sensory approach to our project and have produced systems involved in the sensation of nitric oxide (NO). Originally we set out to develop a bacterial and mammalian hybrid NO-sensing promoter (which we have achieved); we then looked into ways of quantifying the levels of highly reactive and difficult to measure NO within a system, leading to us producing a novel gene regulation system known as the comparator circuit. Throughout the project we went on to look at theoretical alternative approaches to the gene systems we have produced.<br />
<br />
====Overall in our project we have produced 6 sensory BioBricks, 2 BioBricks involved in gene regulation, and have further characterised 4 more BioBricks. All 8 of our BioBricks have been submitted to the registry and the 6 sensory BioBricks have been characterised. Our project consists of three components, read their summaries on this page or click on the images below to see more detailed information including our BioBrick characterisation data.====<br />
<br />
<br />
<br />
{| style="color:#1b2c8a;background-#6C0;" cellpadding="3" cellspacing="1"width="62%" align="center"<br />
|-<br />
|<br />
!align="left"|[[File:NRPNOLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing |300px]]<br />
!align="left"|[[File:NRPCompLogo.png |link=https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit |300px]]<br />
!align="left"|[[File:NRPTheoreticalLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/TheoreticalProjects |300px]]<br />
|-<br />
|<br />
!align="left"|6 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 BBa_K774000] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 BBa_K774001] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 BBa_K774005] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 BBa_K774006] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004]<br />
!align="left"|2 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774002 BBa_K774002] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774003 BBa_K774003]<br />
!align="left"|Theoretical BioBricks: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774100 BBa_K774100]<br />
|-<br />
|<br />
|}<br />
<br />
====Existing iGEM BioBrick parts which we characterised during our project:====<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 PyeaR Promoter BBa_K216005] -- [http://partsregistry.org/Part:BBa_K381001 PyeaR Promoter + GFP BBa_K381001] -- [http://partsregistry.org/Part:BBa_E0420 enhanced Cyan Fluorescent Protein + RBS + Terminators BBa_E0420] -- [http://partsregistry.org/Part:BBa_K081014 Red Fluorescent Protein + RBS + Terminators BBa_K081014]<br />
<br />
----<br />
<br />
[[File:NRPNOLogo.png | centre | link=Team:NRP-UEA-Norwich/NOSensing]]<br />
<br />
<br />
Our main project has resulted in the production of a hybrid bacterial and mammalian promoter optimised for induction by nitric oxide, nitrates and nitrites. We have ligated PyeaR, a known bacterial promoter [http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005] (Cambridge 2009) in the parts registry, with its mammalian counterpart, CArG. The resulting hybrid promoter has been synthesised in two orientations; PyeaR (bacterial, B) upstream of CArG (mammalian, M), nicknamed (B-M); and CArG upstream of PyeaR (M-B). These orientations were submitted to the parts registry as our first two BioBricks.<br />
<br />
Each orientation of the promoter was ligated to enhanced cyan fluorescent protein (eCFP) and red fluorescent protein (RFP) to produce four new BioBricks which have been submitted to the parts registry. These promoter + fluorescent protein BioBricks have been characterised following transformation into ''Escherichia coli'' and induction by potassium nitrate using methods such as flow cytometry, fluorescence-activated cell sorting (FACS) and scanning with a fluorometer. The data from these experiments has proved that our promoter works as we expected it to. <br />
<br />
We have also transfected M-B + eCFP into a human breast cancer cell line, MCF7, and have provided data to show that the system is flexible and can be used in both eukaryotes and prokaryotes. We believe the promoters we have produced have relevant uses in cancer therapeutics, soil fertilisation and detection of emissions from industries such as construction.<br />
<br />
Our 6 BioBricks:<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian/B-M (PyeaR-CArG) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial/M-B (CArG-PyeaR) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP]<br />
<br />
----<br />
<br />
<br />
[[File:NRPCompLogo.png | centre | link=Team:NRP-UEA-Norwich/ComparatorCircuit]]<br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
The lack of specificity of the bacterial promoter, pYeaR, used in the hybrid promoter was a pitfall that was always a concern when it came to accurately sensing nitric oxide. From this potential problem spawned a potential solution; the Comparator Circuit. This pair of BioBricks are designed to specifically bind to each other while ligated to two different promoters of overlapping specificity to result in an integration of the conflicting outputs of the two opposing gene systems.<br />
<br />
<br />
Our system relies on two constructs that interact via complimentary base pair sequences both before and after the ribosome binding site of the reporter protein. The idea being that, when both transcripts are present in the chassis, they would bind together, inhibiting the translation of the reporter proteins.<br />
<br />
<br />
Any imbalance of transcription due to the presence of the substrate of interest results in free mRNA of the gene system that detects that substrate. Crucially, if both promoters detect the same substrates but differ with one extra substrate being detected by one of the promoters, it is this substrate and this substrate only that our system will be able to detect in a simple and quantitative way. <br />
<br />
<br />
Our team have constructed a countercurrent comparator circuit in which the reporter proteins are at the same end of the complimentary region, although a contracurrent system has been theorised. Both systems share a crucial subtractive nature comparable to an analogue computer. We envisage that, should the system be fine-tuned and expanded on, a variety of different business sectors from agriculture to spinoff pharmaceutical companies (<html><font size=2pt><b>Please check out our <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details</b></font></html>) could capitalise on this novel genetic technology.<br />
<br />
<br />
What we have produced is a BioBrick pair that work in harmony, when ligated to promoters of interest and genes of interest, to sequester translation when both mRNA transcripts are present in the cell. The use of quantative tuners with these BioBricks is encouraged to ensure that the transcription rate both gene constructs are equal when both promoters are transcribing at their optimal rate. Although the parts have been submitted to the registry and theoretically characterised, time constraints have meant that further lab-based characterisation could not occur.<br />
<br />
<br />
However, we hope to utilise any free time in our timetables sduring the next semester to characterise the BioBricks further (please see our project proposal), and hope that we will be given a chance to present our further findings at MIT! <br />
<br />
<br />
To conclude, what we have created is a pair of antagonistic BioBricks that turned the pair of mRNAs in which they reside into translational repressor molecules when both are transcribed in tandum within a specific chassis of interest, a new application for mRNA complimentary base pairing within the registry and a project we feel could go very far indeed.<br />
<br />
<br />
----<br />
<br />
<br />
[[File:NRPTheoreticalLogo.png | centre | link=Team:NRP-UEA-Norwich/TheoreticalProjects]]<br />
<br />
<br />
'''Multiplicative circuit'''<br />
<br />
The comparator circuit that we have created integrates two different transcription levels in a negative manner (subtraction) to perform a range of functions a range of different integrations (calculations) are necessary to this end we have also designed a system that would allow one signal (transcription rate) to be divided by the transcription level of another promoter. To achieve this we have used the processes of attenuation and the three loop system (in the trp leader). the system has also been mathematically modeled. To find out more please click the image above. <br />
<br />
<br />
'''Multisensor'''<br />
<br />
There are many groups of chemical species for which there are no current biological techniques for distinguishing between each of these species and quantitatively analysing its concentration. Here we outline a possible approach for solving this problem using non-specific transcription factors and promoters. We use nitrates, nitrites and nitric oxide as our example group. To find more please click the image above.<br />
<br />
<br />
In addition to the two main projects, we have also worked to elaborate on some of the teams earlier ideas during the project.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/ProjectTeam:NRP-UEA-Norwich/Project2012-09-26T23:41:53Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRP}}<br />
<br />
==NRP-UEA-Norwich Project Overview==<br />
<br />
Sensory BioBrick systems have been a large constituent of previous iGEM projects in which teams have combined impressive amounts of logic with limitless creativity in order to produce synthetically engineered organisms with the ability to detect the presence of specific substrates; this was achieved by combining various promoters and reporters to produce novel gene systems of great breadth and depth.<br />
<br />
We too have taken a sensory approach to our project and have produced systems involved in the sensation of nitric oxide (NO). Originally we set out to develop a bacterial and mammalian hybrid NO-sensing promoter (which we have achieved); we then looked into ways of quantifying the levels of highly reactive and difficult to measure NO within a system, leading to us producing a novel gene regulation system known as the comparator circuit. Throughout the project we went on to look at theoretical alternative approaches to the gene systems we have produced.<br />
<br />
====Overall in our project we have produced 6 sensory BioBricks, 2 BioBricks involved in gene regulation, and have further characterised 4 more BioBricks. All 8 of our BioBricks have been submitted to the registry and the 6 sensory BioBricks have been characterised. Our project consists of three components, read their summaries on this page or click on the images below to see more detailed information including our BioBrick characterisation data.====<br />
<br />
<br />
<br />
{| style="color:#1b2c8a;background-#6C0;" cellpadding="3" cellspacing="1"width="62%" align="center"<br />
|-<br />
|<br />
!align="left"|[[File:NRPNOLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing |300px]]<br />
!align="left"|[[File:NRPCompLogo.png |link=https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit |300px]]<br />
!align="left"|[[File:NRPTheoreticalLogo.png|link=https://2012.igem.org/Team:NRP-UEA-Norwich/TheoreticalProjects |300px]]<br />
|-<br />
|<br />
!align="left"|6 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 BBa_K774000] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 BBa_K774001] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 BBa_K774005] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 BBa_K774006] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 BBa_K774004]<br />
!align="left"|2 BioBricks created: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774002 BBa_K774002] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774003 BBa_K774003]<br />
!align="left"|Theoretical BioBricks: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774100 BBa_K774100]<br />
|-<br />
|<br />
|}<br />
<br />
====Existing iGEM BioBrick parts which we characterised during our project:====<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 PyeaR Promoter BBa_K216005] -- [http://partsregistry.org/Part:BBa_K381001 PyeaR Promoter + GFP BBa_K381001] -- [http://partsregistry.org/Part:BBa_E0420 enhanced Cyan Fluorescent Protein + RBS + Terminators BBa_E0420] -- [http://partsregistry.org/Part:BBa_K081014 Red Fluorescent Protein + RBS + Terminators BBa_K081014]<br />
<br />
----<br />
<br />
[[File:NRPNOLogo.png | centre | link=Team:NRP-UEA-Norwich/NOSensing]]<br />
<br />
<br />
Our main project has resulted in the production of a hybrid bacterial and mammalian promoter optimised for induction by nitric oxide, nitrates and nitrites. We have ligated PyeaR, a known bacterial promoter [http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005] (Cambridge 2009) in the parts registry, with its mammalian counterpart, CArG. The resulting hybrid promoter has been synthesised in two orientations; PyeaR (bacterial, B) upstream of CArG (mammalian, M), nicknamed (B-M); and CArG upstream of PyeaR (M-B). These orientations were submitted to the parts registry as our first two BioBricks.<br />
<br />
Each orientation of the promoter was ligated to enhanced cyan fluorescent protein (eCFP) and red fluorescent protein (RFP) to produce four new BioBricks which have been submitted to the parts registry. These promoter + fluorescent protein BioBricks have been characterised following transformation into ''Escherichia coli'' and induction by potassium nitrate using methods such as flow cytometry, fluorescence-activated cell sorting (FACS) and scanning with a fluorometer. The data from these experiments has proved that our promoter works as we expected it to. <br />
<br />
We have also transfected M-B + eCFP into a human breast cancer cell line, MCF7, and have provided data to show that the system is flexible and can be used in both eukaryotes and prokaryotes. We believe the promoters we have produced have relevant uses in cancer therapeutics, soil fertilisation and detection of emissions from industries such as construction.<br />
<br />
Our 6 BioBricks:<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian/B-M (PyeaR-CArG) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial/M-B (CArG-PyeaR) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP]<br />
<br />
----<br />
<br />
<br />
[[File:NRPCompLogo.png | centre | link=Team:NRP-UEA-Norwich/ComparatorCircuit]]<br />
<br />
[[File:Comparator circuit.png | 200px |right | thumbnail | '' '''Figure 3.''' The electrical circuitary of the comparator circuit.'']]<br />
<br />
The lack of specificity of the bacterial promoter, pYeaR, used in the hybrid promoter was a pitfall that was always a concern. From this potential problem spawned a potential solution; the Comparator Circuit.This pair of BioBricks are designed to specifically bind to each other while ligated to two different promoters of overlapping specificity to result in an integration of the conflicting outputs of the two opposing gene systems.<br />
<br />
<br />
Our system relies on two constructs that interact via complimentary base pair sequences both before and after the ribosome binding site of the reporter protein. The idea being that, when both transcripts are present in the chassis, they would bind together, inhibiting the translation of the reporter proteins.<br />
<br />
<br />
Any imbalance of transcription due to the presence of the substrate of interest results in free mRNA of the gene system that detects that substrate. Crucially, if both promoters detect the same substrates but differ with one extra substrate being detected by one of the promoters, it is this substrate and this substrate only that our system will be able to detect in a simple and quantitative way. <br />
<br />
<br />
Our team have constructed a countercurrent comparator circuit in which the reporter proteins are at the same end of the complimentary region, although a contracurrent system has been theorised. Both systems share a crucial subtractive nature comparable to an analogue computer. We envisage that, should the system be fine-tuned and expanded on, a variety of different business sectors from agriculture to spinoff pharmaceutical companies (<html><font size=2pt><b>Please check out our <a href="https://2012.igem.org/Team:NRP-UEA-Norwich/Quanticare">Quanticare</a> page for more details</b></font></html>) could capitalise on this novel genetic technology.<br />
<br />
<br />
What we have produced is a BioBrick pair that work in harmony, when ligated to promoters of interest and genes of interest, to sequester translation when both mRNA transcripts are present in the cell. The use of quantative tuners with these BioBricks is encouraged to ensure that the transcription rate both gene constructs are equal when both promoters are transcribing at their optimal rate. Although the parts have been submitted to the registry and theoretically characterised, time constraints have meant that further lab-based characterisation could not occur.<br />
<br />
<br />
However, we hope to utilise any free time in our timetables sduring the next semester to characterise the BioBricks further (please see our project proposal), and hope that we will be given a chance to present our further findings at MIT! <br />
<br />
<br />
To conclude, what we have created is a pair of antagonistic BioBricks that turned the pair of mRNAs in which they reside into translational repressor molecules when both are transcribed in tandum within a specific chassis of interest, a new application for mRNA complimentary base pairing within the registry and a project we feel could go very far indeed.<br />
<br />
<br />
----<br />
<br />
<br />
[[File:NRPTheoreticalLogo.png | centre | link=Team:NRP-UEA-Norwich/TheoreticalProjects]]<br />
<br />
<br />
'''Multiplicative circuit'''<br />
<br />
The comparator circuit that we have created integrates two different transcription levels in a negative manner (subtraction) to perform a range of functions a range of different integrations (calculations) are necessary to this end we have also designed a system that would allow one signal (transcription rate) to be divided by the transcription level of another promoter. To achieve this we have used the processes of attenuation and the three loop system (in the trp leader). the system has also been mathematically modeled. To find out more please click the image above. <br />
<br />
<br />
'''Multisensor'''<br />
<br />
There are many groups of chemical species for which there are no current biological techniques for distinguishing between each of these species and quantitatively analysing its concentration. Here we outline a possible approach for solving this problem using non-specific transcription factors and promoters. We use nitrates, nitrites and nitric oxide as our example group. To find more please click the image above.<br />
<br />
<br />
In addition to the two main projects, we have also worked to elaborate on some of the teams earlier ideas during the project.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:41:04Z<p>RussellGritton: /* Making future iGEM competitions safer through biosafety engineering */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
<br />
<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed: [https://static.igem.org/mediawiki/2012/2/2e/UEA_GMO_rules.pdf UEA Genetically Modified Organisms Guidelines], [https://static.igem.org/mediawiki/2012/6/65/UEA_statment_of_health_and_saftey_policy.pdf UEA Health and Saftey Policy] and [https://static.igem.org/mediawiki/2012/6/68/UEA_Microbiological_saftey_rules.pdf UEA Microbiological Saftey Rules and Guidelines]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here [http://www.hse.gov.uk/biosafety/gmo/law.htm 'GMOs and the Law'] and [http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp 'The SACGM Compendium of guidance']. All of our projects are in compliance with both university requirements and national regulations.<br />
<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:39:50Z<p>RussellGritton: /* The University of East Anglia's biosafety review board */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
<br />
<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed: [https://static.igem.org/mediawiki/2012/2/2e/UEA_GMO_rules.pdf UEA Genetically Modified Organisms Guidelines], [https://static.igem.org/mediawiki/2012/6/65/UEA_statment_of_health_and_saftey_policy.pdf UEA Health and Saftey Policy] and [https://static.igem.org/mediawiki/2012/6/68/UEA_Microbiological_saftey_rules.pdf UEA Microbiological Saftey Rules and Guidelines]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here [http://www.hse.gov.uk/biosafety/gmo/law.htm 'GMOs and the Law'] and [http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp 'The SACGM Compendium of guidance']. All of our projects are in compliance with both university requirements and national regulations.<br />
<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:39:10Z<p>RussellGritton: /* The University of East Anglia's biosafety review board */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
<br />
<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed: [https://static.igem.org/mediawiki/2012/2/2e/UEA_GMO_rules.pdf UEA Genetically Modified Organisms Guidelines], [https://static.igem.org/mediawiki/2012/6/65/UEA_statment_of_health_and_saftey_policy.pdf UEA Health and Saftey Policy]and[https://static.igem.org/mediawiki/2012/6/68/UEA_Microbiological_saftey_rules.pdf UEA Microbiological Saftey Rules and Guidelines]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here [http://www.hse.gov.uk/biosafety/gmo/law.htm 'GMOs and the Law'], [http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp 'The SACGM Compendium of guidance']. All of our projects are in compliance with both university requirements and national regulations.<br />
<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:38:29Z<p>RussellGritton: /* The University of East Anglia's biosafety review board */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
<br />
<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed:[https://static.igem.org/mediawiki/2012/2/2e/UEA_GMO_rules.pdf UEA Genetically Modified Organisms Guidelines],[https://static.igem.org/mediawiki/2012/6/65/UEA_statment_of_health_and_saftey_policy.pdf UEA Health and Saftey Policy]and[https://static.igem.org/mediawiki/2012/6/68/UEA_Microbiological_saftey_rules.pdf UEA Microbiological Saftey Rules and Guidelines]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here [http://www.hse.gov.uk/biosafety/gmo/law.htm 'GMOs and the Law'], [http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp 'The SACGM Compendium of guidance']. All of our projects are in compliance with both university requirements and national regulations.<br />
<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:34:42Z<p>RussellGritton: /* Safety issues our BioBrick parts could raise: */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
<br />
<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed:[[File:UEA_GMO_rules.pdf|UEA Genetically modified organisms guidelines]],[[File:UEA_statment_of_health_and_saftey_policy.pdf|UEA health and saftey policy]]and[[File:UEA_Microbiological_saftey_rules.pdf|UEA Microbiological saftey rules and guidelines]]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here http://www.hse.gov.uk/biosafety/gmo/law.htm, http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp. All our projects are in compliance with both university requirements and national regulations.<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:33:44Z<p>RussellGritton: /* Environmental safety */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed:[[File:UEA_GMO_rules.pdf|UEA Genetically modified organisms guidelines]],[[File:UEA_statment_of_health_and_saftey_policy.pdf|UEA health and saftey policy]]and[[File:UEA_Microbiological_saftey_rules.pdf|UEA Microbiological saftey rules and guidelines]]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here http://www.hse.gov.uk/biosafety/gmo/law.htm, http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp. All our projects are in compliance with both university requirements and national regulations.<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:33:27Z<p>RussellGritton: /* Public safety */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed:[[File:UEA_GMO_rules.pdf|UEA Genetically modified organisms guidelines]],[[File:UEA_statment_of_health_and_saftey_policy.pdf|UEA health and saftey policy]]and[[File:UEA_Microbiological_saftey_rules.pdf|UEA Microbiological saftey rules and guidelines]]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here http://www.hse.gov.uk/biosafety/gmo/law.htm, http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp. All our projects are in compliance with both university requirements and national regulations.<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/SafetyTeam:NRP-UEA-Norwich/Safety2012-09-26T23:32:55Z<p>RussellGritton: /* Researcher safety */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
=Potential safety issues our project could raise:=<br />
<br />
When designing the experiments the NRP-UEA-Norwich team would undertake they had to consider not only the safety of the researchers and others within the lab, but also the public and the environment. They had to consider carefully the organisms and chemicals they would be using within lab, how they would handle these safely and then dispose of them after to minimize their effect on the environment and the public.<br />
<br />
==Researcher safety==<br />
<br />
[[File:Joy_gloves.jpg |200px|left]]The University of East Anglia already has safety standards that have to be met, which included having basic safety training before starting wet lab work, as well as reading, understanding and signing relevant COSH forms for each potential lab procedure. Therefore, when designing experiments the NRP-UEA-Norwich team considered the risks associated and checked whether the COSH forms signed covered the procedures undertaken. The team also received training from advisors over the first few days of their time in the lab. They learnt what to do in emergencies, where to access the safety manual which is always within the lab, as well as being shown where to dispose of the different chemicals and used equipment. It is also vital that researchers within the university wear personal protective equipment (PPE), including lab coat, gloves and covered shoes. The members also wore protective face shields when using artificial UV light in the darkroom. In accordance with university regulation, the team was prohibited from eating, drinking or smoking in or near the lab. The team enjoyed their lunch well away from the lab, as well as being careful to wash their hands before leaving.<br />
<br />
<br />
The lab facilities were designed to be safe for the team members, providing plenty of space and clean surfaces to work on. The lab was supplied with plastic handheld pipettes, and permanent pens; removing the need for use researchers to ever use their mouths during experimentation e.g. mouth pipettes and labels that require saliva to aid adhesion.<br />
<br />
<br />
The team had originally thought it would be interesting to look at NO levels within "Salmonella", but decided later that this would result in a lot more safety concerns and complications. Therefore, the team decided to only use strains of ''E.coli'' (which they had all used within their university studies before and were allowed to handle within the lab) within our projects. NEB 5-alpha ''E.coli'' was used to characterize existing bio bricks, as well as BL21 pLysS ''E. coli'' cells and Alpha select gold ''E.coli''. ''E.coli'' is a well-studied species of bacteria, with fairly predictable behavior, as well as being non-pathogenic (a bio safety level 1 bacteria). Therefore, all the students were able to use the ''E.coli'' within the university’s second year (category one and two) teaching labs.<br />
<br />
<br />
The hybrid promoter was further characterized by being placed into the mammalian cell MCF7. This was carried out by an experienced member of the team. This caused no concern as the mammalian cells were used and kept under standard laboratory conditions. However, if these cells somehow did make it in to the body they would be recognized as non self cells and consequently destroyed by the immune system. <br />
<br />
<br />
Since the team’s project involves Nitric Oxide (NO) they had to consider the safety of the chemicals involved. It is important to note that all chemicals within the lab are supplied with a safety advice sheet that has been generated by the company it was manufactured by. NO has been classed as a dangerous gas which can be directly toxic to blood, lungs, pancreas and nervous system. Therefore, caution was taken when using NO (within the form of potassium nitrate) within the lab. <br />
<br />
<br />
[[File:Protection_of_ears.jpg |300px|right]]<br />
<br />
<br />
Another safety concern was the use of Ethidium Bromide (EtBr) during the agrose gel staining. Ethidium Bromide is a known carcinogen, therefore the team took care to follow the university's specific guidelines on working with carcinogens ([https://static.igem.org/mediawiki/2012/4/4b/UEA_Rules_for_working_with_carcinogens_in_laboratories.pdf Rules For Working With Carcinogens]), which include the use of protective equipment, gloves and the appropriate disposal of waste in order to minimize researcher exposure.<br />
<br />
<br />
In the preliminary discussions in which the experiments the team would carry out were discussed, careful consideration was taken to choose appropriate equipment. They considered the equipment’s safety to the researcher, as well as the practicality and quality of results gained. The team members used most of the discussed equipment and procedures already used during their first two years of their undergraduate degree. However, there were a few procedures such as NanoDrop spectrophotometery and fluorometery that had not been used by the team before. Therefore the team members received training from experienced members of staff in order to avoid accidents that could damage the researchers or the machine itself. Finally, there were always more experienced scientists present within the lab to advise the team further on how to use equipment safely.<br />
<br />
==Public safety==<br />
<br />
The team always removed their lab coats and gloves, as well as washed their hands, before leaving the lab. This reduced the chance of bringing harmful bacteria or substances outside the laboratory environment. The team also took care to keep the laboratory windows shut, in order to reduce exposure to the outside world.<br />
<br />
All contaminated waste was autoclaved on site before being collected and disposed of under controlled conditions, or for cleaning for reuse.<br />
<br />
The team felt that it was important that the public were given the opportunity to understand and ask about procedures used in the laboratory, so they could form informed opinions about the safety and ethical implications of the teams work. The team took the opportunity at their public engagement event to explain the procedures they used in full detail.<br />
<br />
At this public engagement event the team took transformed ''E.coli'' samples to the venues and public safety was a priority. Therefore, the plates were maintained in a UV box behind a sheet of plastic, limiting public physical interaction with the laboratory specimens. These plates were also sealed extensively using parafilm and were kept well away form any part of the venue holding or serving food products.<br />
<br />
==Environmental safety==<br />
<br />
[[File:Rabbits_at_UEA.png| 300px||left ]]Data safety sheets that come with chemicals explain how to dispose of the chemicals correctly. These guidelines were adhered to rigidly since the local water providing company, Anglia water, carry out routine random samples on what is poured down the sink drains, and if a certain volume (volume is unknown to university, which encourages them not to put any waste down the sink) of listed chemicals are found, then the university is fined.<br />
<br />
There is a very low risk of damage to the environment as all bacteria will be stored safely within the lab. However, if the bacteria was to unexpectedly be released, there is very little threat on the environment, since the bacteria are standard autotrophic laboratory stains and therefore unlikely to survive. The ''E.coli'' stains used are also unlikely to replicate and therefore spread. Finally, due to using plasmids with a narrow host range horizontal gene transfer between bacteria within the environment is also very unlikely, as plasmids would not replicate if transferred to other organisms.<br />
<br />
<br />
<br />
=Safety issues our BioBrick parts could raise:=<br />
<br />
The first BioBricks made by the NRP-UEA iGEM team are hybrid promoters consisting of a bacterial promoter (B) fused to a mammalian promoter (M); this has been carried out in two orientations, B upstream from M (B-M), and M upstream from B (M-B). As this is a new concept the BioBricks safety concerns arise, due to the lack of knowledge surrounding how the promoters will interact with one another. The team has not submitted any issues to the registry thus far as no specific issues have arisen, there are no specific concerns regarding the DNA sequence and both promoters are individually well characterised; however the team remains vigilant and understands that with unknown BioBricks such as B-M and M-B there is always the chance for something unexpected to occur.<br />
[[File:MB.png|200px|right]]<br />
[[File:BM.png|200px|right]]<br />
<br />
The team has also synthesised a second new construct known as the comparator circuit, which again is not well characterised. Through its intended use is as a gene regulation component a risk arises in that the comparator circuit could possibly join to unintended, functional genes and affect their regulation. As no specific issues have arisen yet no issues have been added to the registry, however upon characterisation of the BioBricks any issues that do arise will immediately be included with the part.<br />
<br />
Throughout the project the team handled all safety issues by following and complying with both government safety standards as well as university safety standards. The team has also inserted the BioBricks into narrow host-range plasmids (as produced by iGEM), thus reducing the chances of horizontal gene transfer. BioBricks of B-M, M-B, and the comparator circuit, were all transformed into standard BSL1 laboratory autotrophic strains of ''E. coli'' and are therefore unlikely to survive outside of the laboratory.<br />
<br />
Future iGEM teams could increase their safety by connecting the B-M and M-B BioBricks to a suicide gene. This would mean that in the unlikely event of the transformed cells being released into the environment and unintended bacteria taking up the BioBrick-containing plasmid, the unintended bacteria would be destroyed and further spread of the hybrid promoter BioBricks would be prevented.<br />
<br />
=The University of East Anglia's biosafety review board=<br />
<br />
UEA takes both staff and student safety very seriously and has its own health and safety policies and biosaftey rules that must be followed:[[File:UEA_GMO_rules.pdf|UEA Genetically modified organisms guidelines]],[[File:UEA_statment_of_health_and_saftey_policy.pdf|UEA health and saftey policy]]and[[File:UEA_Microbiological_saftey_rules.pdf|UEA Microbiological saftey rules and guidelines]]. These have been created to follow the guidelines and biosafty rules that have to be considered in the United Kingdom as seen here http://www.hse.gov.uk/biosafety/gmo/law.htm, http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp. All our projects are in compliance with both university requirements and national regulations.<br />
<br />
The university also has its own biosafety committee in which the responsibilities of different aspects of safety are split between departments and staff. Dr Andrew Hemmings is the BIO safety officer at UEA and spoke to members of the team about the project and discussed whether our COSH forms covered the processes carried out. The team then discussed their project further with other members of staff including Dr Mark Coleman, university GMO safety officer, and Dr Gabriella Kelemen, university microbial safety officer. <br />
<br />
When speaking to members of the UEA biosafty team there was no concern with the processes being carried out and the bacteria being used. When the team spoke to GMO safety officer, Dr Mark Coleman, about the safety of the BioBricks they intended to create he said "there is a highly improbable chance this project poses a significant risk to either human health or the environment".<br />
<br />
=Making future iGEM competitions safer through biosafety engineering=<br />
<br />
Although the team started looking at NO, the dangers of unintentional exposure to NO soon became clear. Therefore, the team would hope that through using systems similar to our comparator sensor, NO sensors will be produced that are in the form of a sheet of paper and can be placed on the wall of labs, and buildings ect, to indicate excess levels of NO visually. <br />
<br />
The team feel that future iGEM teams could aim to reduce the chance of horizontal gene transfer further by reducing the adherence between the transformed cells and others, or via the prevention of transformations without inducing competency. This would be a procedure to prevent some cases of antibiotic genes being passed through between bacterial species. However, a variety of other safeguards could also be deployed to prevent this.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/QuanticareTeam:NRP-UEA-Norwich/Quanticare2012-09-26T23:21:47Z<p>RussellGritton: /* Ethics of Quanticare and Cura */</p>
<hr />
<div>{{UEANRPQuanticare}}<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table><br><b>The NRP UEA iGEM team would like to thank the Biochemical Society for funding of our film, and Amy Congdon for direction and production of our film.</b><br />
</center></html><br />
<br />
==Quanticare & The ''Cura'' Tattoo==<br />
<br />
As part of our human practices we have produced a film in conjunction with our artist [http://www.amycongdon.com/#!mainPage Amy Congdon] which explores a futuristic world where synthetic biology is an aspect of everyday life. As we explored the idea of synthetic biology in the future we developed the idea of "Quanticare", a company created on the back of synthetic biology's rise to prominence. As the project within the labs continued and the idea of the [https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit Comparator Circuit] and the [https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing Flexible Bacterial/Mammalian Hybrid Promoter] came about we looked at how this new idea of cellular computing could be used within the Quanticare concept, leading to the idea of the living tattoo, ''Cura''.<br />
<br />
<br />
We wanted to produce something that would spark the imagination of the wider public and truly show the future power and potential of synthetic biology, and we believe that ''Cura'' and Quanticare have achieved those aims. With ''Cura'' we have developed an original idea of a tattoo not drawn with ink, but instead drawn with immortalised mammalian cells containing different BioBricks. The concept involves each shape of the tattoo containing cells with specific BioBricks that sense a particular chemical within the body, and report on any particular changes on that chemical's levels. <br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
<br />
The comparator circuit project involves multiple promoters that are sensitive to similar chemicals, a different reporter/effector enzyme for each promoter, and a range of specially designed DNA leaders that go between the promoter and reporter of each gene system. The idea involves the specially designed DNA leaders being complementary to one another and resulting in DNA annealing which covers the ribosome binding site; this means that mRNAs transcribed following activation of the promoters will cancel one another out and fail to be translated. If there is an imbalance of substrates one set of mRNAs would be produced in excess of the others as this set would not anneal to one another, therefore resulting in translation and expression of the associated protein. This is demonstrated in ''Figure 1.'' where two sets of mRNA have been produced, one with an RFP reporter and one with a CFP reporter; due to an imbalance in substrates four lots of the RFP-associated mRNA have been produced, while only two lots of the CFP-associated mRNAs have been produced. As the image shows the two CFP-associated mRNAs have annealed with two of the RFP-associated mRNAs, blocking the RBS and preventing translation, however due to the imbalance there are two RFP-associated mRNAs left to be translated and thus RFP would be expressed in this situation.<br />
<br />
<br />
Similar comparator circuit set-ups would be used in the ''Cura'' tattoo. For instance in the case of Type I Diabetes one promoter would sense blood sugar while another would sense insulin; if the blood sugar outweighed the insulin, more of the mRNA with the blood sugar promoter would be produced and thus an effector enzyme for the production of insulin could be included in this mRNA in order to increase insulin production. If insulin production becomes too high the imbalance would shift so more of the mRNA with the insulin-sensing promoter was produced, and an effector enzyme to break down excess insulin could be translated from this mRNA strand in order to normalise insulin levels. A system such as this, as well as others (which simply report changes through fluorescent proteins), are explored in Figure 2.<br />
<br />
<br />
[[File:NRPCura.png | center | 950 px | thumbnail | '''Figure 2.''' ''Cura'' is Quanticare's revolutionary healthcare solution, a tattoo made of immortalised mammalian cells containing different biobricks that have been specifically tailored to monitor and respond to changes in the body. Different areas of the tattoo correspond to different areas of the body, and using biological technology developed by the Norwich Research Park and University of East Anglia 2012 iGEM Team can react automatically to many unwanted changes, as well as fluorescing to alert the user of any problems.]]<br />
<br />
<br />
We have also looked at therapeutic strategies within Quanticare and how the tattoo may be analysed. As part of the film we looked at the idea of a scanner to complement the ''Cura'' tattoo, which will scan the tattoo's fluorescence patterns and feed through to a specialised printer designed to produce a bacterial pill optimised to treat the conditions reported by the tattoo. These bacteria would contain biobricks to produce drugs/enzymes as well as targeting mechanisms in order to find the site of a problem and correct mistakes being made; they could also be used to help swamp the likes of cancerous tumours and pump the tumour with drugs/apoptosis-inducing factors such as nitric oxide in order to help fight cancer; a mocked-up ''in vitro'' video of this occurring is included in the film. <br />
<br />
<br />
As part of a marketing strategy to promote Quanticare and our film we have produced transferable tattoo versions of ''Cura'' which we will be handing out at the European Jamboree for other competitors, judges and interested parties to enjoy. We believe this will be an interactive and exciting approach to promoting our project and our ideas on the future of synthetic biology.<br />
<br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 before the wiki freeze:<br />
<br />
<br />
. Blogged about on Biochemical Society blog <br />
<br />
<br />
. On UEA news pages <br />
<br />
<br />
. Mentioned in tweet from the BBSRC <br />
<br />
<br />
. Many likes and tweets<br />
<br />
<br />
. Some quotes;<br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist <br />
<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
<br />
===Ethics of Quanticare and ''Cura''===<br />
<br />
Through the film we have also explored the ethics of both synthetic biology healthcare solutions as well as synthetic biology as a whole. We have envisaged a world where public protests regarding synthetic biology have occurred, leading to companies such as Quanticare adopting stringent safety methods and testing in order to help calm public unease. We have also looked at the ethics of a tattoo and tattooing with immortalised cells, in particular the obvious risk of adding foreign cells to a body and public opinion on tattooing. We believe that while there is definite opposition, ''Cura'' is still a realistic future product provided the appropriate safety concerns are addressed.<br />
<br />
<br />
As part of our human practices we ran a synthetic biology-based event at a local events centre, the Norwich Forum; at the event we debuted the video and looked to members of the public to give their opinions on the concept as well as a future involving synthetic biology. Many members of the public were extremely interested in the idea and some even asked if it was available for them to tattoo ''Cura'' onto themselves that day! However, there were also many concerns raised by members of the public which we were able to take on board and discuss with them. These concerns included:<br />
<br />
<br />
. Whether the immortalised cells could result in a cancer forming<br />
<br />
. Tattooing being frowned-upon by certain faiths and areas of society<br />
<br />
. Harmful affects of the cells (e.g. over-production of insulin)<br />
<br />
. Faults in the tattoo systems <br />
<br />
. Encouraging members of the public to rely on the tattoo rather than using healthcare professionals such as doctors<br />
<br />
<br />
We have recognised all of these concerns as valid and appropriate points, and have spoken with the members of the public raising them to discuss how they feel the points should be addressed. Solutions to some of the problems we have investigated include:<br />
<br />
<br />
. Adding a suicide gene in all cells should they begin to divide uncontrollably<br />
<br />
. Careful tuning and testing of all BioBrick systems to prevent over-production of certain products and to troubleshoot any faults<br />
<br />
. In figure 2 some of the strands of the tattoo suggest consulting a doctor, and ''Cura'' is encouraged as both a healthcare solution and an aid to prevent reliance on ''Cura''<br />
<br />
<br />
Overall while there are many ethical issues to be addressed we feel that conversing with the public on these issues and gauging their thoughts is an integral part to moving forward with this concept movie and helping to bring synthetic biology to the forefront of society. We encourage any and all feedback on the film and ''Cura'' as a concept to help improve how synthetic biology is understood and presented.<br />
<br />
<br />
We hope that Quanticare, ''Cura'' and our film are enjoyed by everyone who views them and that they provoke the debates and thoughts we designed the concept to.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/QuanticareTeam:NRP-UEA-Norwich/Quanticare2012-09-26T23:08:04Z<p>RussellGritton: /* Quanticare & The Cura Tattoo */</p>
<hr />
<div>{{UEANRPQuanticare}}<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table><br><b>The NRP UEA iGEM team would like to thank the Biochemical Society for funding of our film, and Amy Congdon for direction and production of our film.</b><br />
</center></html><br />
<br />
==Quanticare & The ''Cura'' Tattoo==<br />
<br />
As part of our human practices we have produced a film in conjunction with our artist [http://www.amycongdon.com/#!mainPage Amy Congdon] which explores a futuristic world where synthetic biology is an aspect of everyday life. As we explored the idea of synthetic biology in the future we developed the idea of "Quanticare", a company created on the back of synthetic biology's rise to prominence. As the project within the labs continued and the idea of the [https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit Comparator Circuit] and the [https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing Flexible Bacterial/Mammalian Hybrid Promoter] came about we looked at how this new idea of cellular computing could be used within the Quanticare concept, leading to the idea of the living tattoo, ''Cura''.<br />
<br />
<br />
We wanted to produce something that would spark the imagination of the wider public and truly show the future power and potential of synthetic biology, and we believe that ''Cura'' and Quanticare have achieved those aims. With ''Cura'' we have developed an original idea of a tattoo not drawn with ink, but instead drawn with immortalised mammalian cells containing different BioBricks. The concept involves each shape of the tattoo containing cells with specific BioBricks that sense a particular chemical within the body, and report on any particular changes on that chemical's levels. <br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
<br />
The comparator circuit project involves multiple promoters that are sensitive to similar chemicals, a different reporter/effector enzyme for each promoter, and a range of specially designed DNA leaders that go between the promoter and reporter of each gene system. The idea involves the specially designed DNA leaders being complementary to one another and resulting in DNA annealing which covers the ribosome binding site; this means that mRNAs transcribed following activation of the promoters will cancel one another out and fail to be translated. If there is an imbalance of substrates one set of mRNAs would be produced in excess of the others as this set would not anneal to one another, therefore resulting in translation and expression of the associated protein. This is demonstrated in ''Figure 1.'' where two sets of mRNA have been produced, one with an RFP reporter and one with a CFP reporter; due to an imbalance in substrates four lots of the RFP-associated mRNA have been produced, while only two lots of the CFP-associated mRNAs have been produced. As the image shows the two CFP-associated mRNAs have annealed with two of the RFP-associated mRNAs, blocking the RBS and preventing translation, however due to the imbalance there are two RFP-associated mRNAs left to be translated and thus RFP would be expressed in this situation.<br />
<br />
<br />
Similar comparator circuit set-ups would be used in the ''Cura'' tattoo. For instance in the case of Type I Diabetes one promoter would sense blood sugar while another would sense insulin; if the blood sugar outweighed the insulin, more of the mRNA with the blood sugar promoter would be produced and thus an effector enzyme for the production of insulin could be included in this mRNA in order to increase insulin production. If insulin production becomes too high the imbalance would shift so more of the mRNA with the insulin-sensing promoter was produced, and an effector enzyme to break down excess insulin could be translated from this mRNA strand in order to normalise insulin levels. A system such as this, as well as others (which simply report changes through fluorescent proteins), are explored in Figure 2.<br />
<br />
<br />
[[File:NRPCura.png | center | 950 px | thumbnail | '''Figure 2.''' ''Cura'' is Quanticare's revolutionary healthcare solution, a tattoo made of immortalised mammalian cells containing different biobricks that have been specifically tailored to monitor and respond to changes in the body. Different areas of the tattoo correspond to different areas of the body, and using biological technology developed by the Norwich Research Park and University of East Anglia 2012 iGEM Team can react automatically to many unwanted changes, as well as fluorescing to alert the user of any problems.]]<br />
<br />
<br />
We have also looked at therapeutic strategies within Quanticare and how the tattoo may be analysed. As part of the film we looked at the idea of a scanner to complement the ''Cura'' tattoo, which will scan the tattoo's fluorescence patterns and feed through to a specialised printer designed to produce a bacterial pill optimised to treat the conditions reported by the tattoo. These bacteria would contain biobricks to produce drugs/enzymes as well as targeting mechanisms in order to find the site of a problem and correct mistakes being made; they could also be used to help swamp the likes of cancerous tumours and pump the tumour with drugs/apoptosis-inducing factors such as nitric oxide in order to help fight cancer; a mocked-up ''in vitro'' video of this occurring is included in the film. <br />
<br />
<br />
As part of a marketing strategy to promote Quanticare and our film we have produced transferable tattoo versions of ''Cura'' which we will be handing out at the European Jamboree for other competitors, judges and interested parties to enjoy. We believe this will be an interactive and exciting approach to promoting our project and our ideas on the future of synthetic biology.<br />
<br />
<br />
Some initial feedback, the Quanticare video was launched less than 24 before the wiki freeze:<br />
<br />
<br />
. Blogged about on Biochemical Society blog <br />
<br />
<br />
. On UEA news pages <br />
<br />
<br />
. Mentioned in tweet from the BBSRC <br />
<br />
<br />
. Many likes and tweets<br />
<br />
<br />
. Some quotes;<br />
<br />
"Inspiring and Innovative" Global Strategist and Author @DeanneLawrence <br />
<br />
<br />
"Great project. Good luck!" @SocratesLogos Synthetic Biologist <br />
<br />
<br />
"Sooooooo cool!" Kimberley Hirst Jones UEA-JIC_Norwich iGEM 2011 team member<br />
<br />
<br />
===Ethics of Quanticare and ''Cura''===<br />
<br />
Through the film we have also explored the ethics of both synthetic biology healthcare solutions as well as synthetic biology as a whole. We have envisaged a world where public protests regarding synthetic biology have occurred, leading to companies such as Quanticare adopting stringent safety methods and testing in order to help calm public unease. We have also looked at the ethics of a tattoo and tattooing with immortalised cells, in particular the obvious risk of adding foreign cells to a body and public opinion on tattooing. We believe that while there is definite opposition, ''Cura'' is still a realistic future product provided the appropriate safety concerns are addressed (e.g. including a suicide gene in all cells should they begin to divide uncontrollably).<br />
<br />
<br />
As part of our human practices we ran a synthetic biology-based event at a local events centre, the Norwich Forum; at the event we debuted the video and looked to members of the public to give their opinions on the concept as well as a future involving synthetic biology. Many members of the public were extremely interested in the idea and some even asked if it was available for them to tattoo ''Cura'' onto themselves that day! However, there were also many concerns raised by members of the public which we were able to take on board and discuss with them. These concerns included:<br />
<br />
<br />
. Whether the immortalised cells could result in a cancer forming<br />
<br />
. Tattooing being frowned-upon by certain faiths and areas of society<br />
<br />
. Harmful affects of the cells (e.g. over-production of insulin)<br />
<br />
. Faults in the tattoo systems <br />
<br />
. Encouraging members of the public to rely on the tattoo rather than using healthcare professionals such as doctors<br />
<br />
<br />
We have recognised all of these concerns as valid and appropriate points, and have spoken with the members of the public raising them to discuss how they feel the points should be addressed. Sol</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/QuanticareTeam:NRP-UEA-Norwich/Quanticare2012-09-26T23:00:50Z<p>RussellGritton: /* Ethics of Quanticare and Cura */</p>
<hr />
<div>{{UEANRPQuanticare}}<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table><br><b>The NRP UEA iGEM team would like to thank the Biochemical Society for funding of our film, and Amy Congdon for direction and production of our film.</b><br />
</center></html><br />
<br />
==Quanticare & The ''Cura'' Tattoo==<br />
<br />
As part of our human practices we have produced a film in conjunction with our artist [http://www.amycongdon.com/#!mainPage Amy Congdon] which explores a futuristic world where synthetic biology is an aspect of everyday life. As we explored the idea of synthetic biology in the future we developed the idea of "Quanticare", a company created on the back of synthetic biology's rise to prominence. As the project within the labs continued and the idea of the [https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit Comparator Circuit] and the [https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing Flexible Bacterial/Mammalian Hybrid Promoter] came about we looked at how this new idea of cellular computing could be used within the Quanticare concept, leading to the idea of the living tattoo, ''Cura''.<br />
<br />
<br />
We wanted to produce something that would spark the imagination of the wider public and truly show the future power and potential of synthetic biology, and we believe that ''Cura'' and Quanticare have achieved those aims. With ''Cura'' we have developed an original idea of a tattoo not drawn with ink, but instead drawn with immortalised mammalian cells containing different BioBricks. The concept involves each shape of the tattoo containing cells with specific BioBricks that sense a particular chemical within the body, and report on any particular changes on that chemical's levels. <br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
<br />
The comparator circuit project involves multiple promoters that are sensitive to similar chemicals, a different reporter/effector enzyme for each promoter, and a range of specially designed DNA leaders that go between the promoter and reporter of each gene system. The idea involves the specially designed DNA leaders being complementary to one another and resulting in DNA annealing which covers the ribosome binding site; this means that mRNAs transcribed following activation of the promoters will cancel one another out and fail to be translated. If there is an imbalance of substrates one set of mRNAs would be produced in excess of the others as this set would not anneal to one another, therefore resulting in translation and expression of the associated protein. This is demonstrated in ''Figure 1.'' where two sets of mRNA have been produced, one with an RFP reporter and one with a CFP reporter; due to an imbalance in substrates four lots of the RFP-associated mRNA have been produced, while only two lots of the CFP-associated mRNAs have been produced. As the image shows the two CFP-associated mRNAs have annealed with two of the RFP-associated mRNAs, blocking the RBS and preventing translation, however due to the imbalance there are two RFP-associated mRNAs left to be translated and thus RFP would be expressed in this situation.<br />
<br />
<br />
Similar comparator circuit set-ups would be used in the ''Cura'' tattoo. For instance in the case of Type I Diabetes one promoter would sense blood sugar while another would sense insulin; if the blood sugar outweighed the insulin, more of the mRNA with the blood sugar promoter would be produced and thus an effector enzyme for the production of insulin could be included in this mRNA in order to increase insulin production. If insulin production becomes too high the imbalance would shift so more of the mRNA with the insulin-sensing promoter was produced, and an effector enzyme to break down excess insulin could be translated from this mRNA strand in order to normalise insulin levels. A system such as this, as well as others (which simply report changes through fluorescent proteins), are explored in Figure 2.<br />
<br />
<br />
[[File:NRPCura.png | center | 950 px | thumbnail | '''Figure 2.''' ''Cura'' is Quanticare's revolutionary healthcare solution, a tattoo made of immortalised mammalian cells containing different biobricks that have been specifically tailored to monitor and respond to changes in the body. Different areas of the tattoo correspond to different areas of the body, and using biological technology developed by the Norwich Research Park and University of East Anglia 2012 iGEM Team can react automatically to many unwanted changes, as well as fluorescing to alert the user of any problems.]]<br />
<br />
<br />
We have also looked at therapeutic strategies within Quanticare and how the tattoo may be analysed. As part of the film we looked at the idea of a scanner to complement the ''Cura'' tattoo, which will scan the tattoo's fluorescence patterns and feed through to a specialised printer designed to produce a bacterial pill optimised to treat the conditions reported by the tattoo. These bacteria would contain biobricks to produce drugs/enzymes as well as targeting mechanisms in order to find the site of a problem and correct mistakes being made; they could also be used to help swamp the likes of cancerous tumours and pump the tumour with drugs/apoptosis-inducing factors such as nitric oxide in order to help fight cancer; a mocked-up ''in vitro'' video of this occurring is included in the film. <br />
<br />
<br />
As part of a marketing strategy to promote Quanticare and our film we have produced transferable tattoo versions of ''Cura'' which we will be handing out at the European Jamboree for other competitors, judges and interested parties to enjoy. We believe this will be an interactive and exciting approach to promoting our project and our ideas on the future of synthetic biology.<br />
<br />
===Ethics of Quanticare and ''Cura''===<br />
<br />
Through the film we have also explored the ethics of both synthetic biology healthcare solutions as well as synthetic biology as a whole. We have envisaged a world where public protests regarding synthetic biology have occurred, leading to companies such as Quanticare adopting stringent safety methods and testing in order to help calm public unease. We have also looked at the ethics of a tattoo and tattooing with immortalised cells, in particular the obvious risk of adding foreign cells to a body and public opinion on tattooing. We believe that while there is definite opposition, ''Cura'' is still a realistic future product provided the appropriate safety concerns are addressed (e.g. including a suicide gene in all cells should they begin to divide uncontrollably).<br />
<br />
<br />
As part of our human practices we ran a synthetic biology-based event at a local events centre, the Norwich Forum; at the event we debuted the video and looked to members of the public to give their opinions on the concept as well as a future involving synthetic biology. Many members of the public were extremely interested in the idea and some even asked if it was available for them to tattoo ''Cura'' onto themselves that day! However, there were also many concerns raised by members of the public which we were able to take on board and discuss with them. These concerns included:<br />
<br />
<br />
. Whether the immortalised cells could result in a cancer forming<br />
<br />
. Tattooing being frowned-upon by certain faiths and areas of society<br />
<br />
. Harmful affects of the cells (e.g. over-production of insulin)<br />
<br />
. Faults in the tattoo systems <br />
<br />
. Encouraging members of the public to rely on the tattoo rather than using healthcare professionals such as doctors<br />
<br />
<br />
We have recognised all of these concerns as valid and appropriate points, and have spoken with the members of the public raising them to discuss how they feel the points should be addressed. Sol</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/QuanticareTeam:NRP-UEA-Norwich/Quanticare2012-09-26T22:51:37Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRPQuanticare}}<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table><br><b>The NRP UEA iGEM team would like to thank the Biochemical Society for funding of our film, and Amy Congdon for direction and production of our film.</b><br />
</center></html><br />
<br />
==Quanticare & The ''Cura'' Tattoo==<br />
<br />
As part of our human practices we have produced a film in conjunction with our artist [http://www.amycongdon.com/#!mainPage Amy Congdon] which explores a futuristic world where synthetic biology is an aspect of everyday life. As we explored the idea of synthetic biology in the future we developed the idea of "Quanticare", a company created on the back of synthetic biology's rise to prominence. As the project within the labs continued and the idea of the [https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit Comparator Circuit] and the [https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing Flexible Bacterial/Mammalian Hybrid Promoter] came about we looked at how this new idea of cellular computing could be used within the Quanticare concept, leading to the idea of the living tattoo, ''Cura''.<br />
<br />
<br />
We wanted to produce something that would spark the imagination of the wider public and truly show the future power and potential of synthetic biology, and we believe that ''Cura'' and Quanticare have achieved those aims. With ''Cura'' we have developed an original idea of a tattoo not drawn with ink, but instead drawn with immortalised mammalian cells containing different BioBricks. The concept involves each shape of the tattoo containing cells with specific BioBricks that sense a particular chemical within the body, and report on any particular changes on that chemical's levels. <br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
<br />
The comparator circuit project involves multiple promoters that are sensitive to similar chemicals, a different reporter/effector enzyme for each promoter, and a range of specially designed DNA leaders that go between the promoter and reporter of each gene system. The idea involves the specially designed DNA leaders being complementary to one another and resulting in DNA annealing which covers the ribosome binding site; this means that mRNAs transcribed following activation of the promoters will cancel one another out and fail to be translated. If there is an imbalance of substrates one set of mRNAs would be produced in excess of the others as this set would not anneal to one another, therefore resulting in translation and expression of the associated protein. This is demonstrated in ''Figure 1.'' where two sets of mRNA have been produced, one with an RFP reporter and one with a CFP reporter; due to an imbalance in substrates four lots of the RFP-associated mRNA have been produced, while only two lots of the CFP-associated mRNAs have been produced. As the image shows the two CFP-associated mRNAs have annealed with two of the RFP-associated mRNAs, blocking the RBS and preventing translation, however due to the imbalance there are two RFP-associated mRNAs left to be translated and thus RFP would be expressed in this situation.<br />
<br />
<br />
Similar comparator circuit set-ups would be used in the ''Cura'' tattoo. For instance in the case of Type I Diabetes one promoter would sense blood sugar while another would sense insulin; if the blood sugar outweighed the insulin, more of the mRNA with the blood sugar promoter would be produced and thus an effector enzyme for the production of insulin could be included in this mRNA in order to increase insulin production. If insulin production becomes too high the imbalance would shift so more of the mRNA with the insulin-sensing promoter was produced, and an effector enzyme to break down excess insulin could be translated from this mRNA strand in order to normalise insulin levels. A system such as this, as well as others (which simply report changes through fluorescent proteins), are explored in Figure 2.<br />
<br />
<br />
[[File:NRPCura.png | center | 950 px | thumbnail | '''Figure 2.''' ''Cura'' is Quanticare's revolutionary healthcare solution, a tattoo made of immortalised mammalian cells containing different biobricks that have been specifically tailored to monitor and respond to changes in the body. Different areas of the tattoo correspond to different areas of the body, and using biological technology developed by the Norwich Research Park and University of East Anglia 2012 iGEM Team can react automatically to many unwanted changes, as well as fluorescing to alert the user of any problems.]]<br />
<br />
<br />
We have also looked at therapeutic strategies within Quanticare and how the tattoo may be analysed. As part of the film we looked at the idea of a scanner to complement the ''Cura'' tattoo, which will scan the tattoo's fluorescence patterns and feed through to a specialised printer designed to produce a bacterial pill optimised to treat the conditions reported by the tattoo. These bacteria would contain biobricks to produce drugs/enzymes as well as targeting mechanisms in order to find the site of a problem and correct mistakes being made; they could also be used to help swamp the likes of cancerous tumours and pump the tumour with drugs/apoptosis-inducing factors such as nitric oxide in order to help fight cancer; a mocked-up ''in vitro'' video of this occurring is included in the film. <br />
<br />
<br />
As part of a marketing strategy to promote Quanticare and our film we have produced transferable tattoo versions of ''Cura'' which we will be handing out at the European Jamboree for other competitors, judges and interested parties to enjoy. We believe this will be an interactive and exciting approach to promoting our project and our ideas on the future of synthetic biology.<br />
<br />
===Ethics of Quanticare and ''Cura''===</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/QuanticareTeam:NRP-UEA-Norwich/Quanticare2012-09-26T22:45:41Z<p>RussellGritton: </p>
<hr />
<div>{{UEANRPQuanticare}}<br />
<br />
<html><center><table width=100% cellpadding=0 cellspacing=0 align=center><tr><td valign=absmiddle align=center><iframe width="560" height="315" src="http://www.youtube.com/embed/StNFePmymbc" frameborder="0" allowfullscreen></iframe></td></tr></table><br><b>The NRP UEA iGEM team would like to thank the Biochemical Society for funding of our film, and Amy Congdon for direction and production of our film.</b><br />
</center></html><br />
<br />
==Quanticare & The ''Cura'' Tattoo==<br />
<br />
As part of our human practices we have produced a film in conjunction with our artist [http://www.amycongdon.com/#!mainPage Amy Congdon] which explores a futuristic world where synthetic biology is an aspect of everyday life. As we explored the idea of synthetic biology in the future we developed the idea of "Quanticare", a company created on the back of synthetic biology's rise to prominence. As the project within the labs continued and the idea of the [https://2012.igem.org/Team:NRP-UEA-Norwich/ComparatorCircuit Comparator Circuit] and the [https://2012.igem.org/Team:NRP-UEA-Norwich/NOSensing Flexible Bacterial/Mammalian Hybrid Promoter] came about we looked at how this new idea of cellular computing could be used within the Quanticare concept, leading to the idea of the living tattoo, ''Cura''.<br />
<br />
<br />
We wanted to produce something that would spark the imagination of the wider public and truly show the future power and potential of synthetic biology, and we believe that ''Cura'' and Quanticare have achieved those aims. With ''Cura'' we have developed an original idea of a tattoo not drawn with ink, but instead drawn with immortalised mammalian cells containing different BioBricks. The concept involves each shape of the tattoo containing cells with specific BioBricks that sense a particular chemical within the body, and report on any particular changes on that chemical's levels. <br />
<br />
[[File:QuanticareComparator.png | right | 250 px | thumbnail | '''Figure 1.''' A graphic showing the result of the comparator circuit; mRNAs transcribed following activation of two promoters have cancelled one another out, leaving only the overhang representative to the imbalance in substrate chemicals to be translated and expressed.]]<br />
<br />
<br />
The comparator circuit project involves multiple promoters that are sensitive to similar chemicals, a different reporter/effector enzyme for each promoter, and a range of specially designed DNA leaders that go between the promoter and reporter of each gene system. The idea involves the specially designed DNA leaders being complementary to one another and resulting in DNA annealing which covers the ribosome binding site; this means that mRNAs transcribed following activation of the promoters will cancel one another out and fail to be translated. If there is an imbalance of substrates one set of mRNAs would be produced in excess of the others as this set would not anneal to one another, therefore resulting in translation and expression of the associated protein. This is demonstrated in ''Figure 1.'' where two sets of mRNA have been produced, one with an RFP reporter and one with a CFP reporter; due to an imbalance in substrates four lots of the RFP-associated mRNA have been produced, while only two lots of the CFP-associated mRNAs have been produced. As the image shows the two CFP-associated mRNAs have annealed with two of the RFP-associated mRNAs, blocking the RBS and preventing translation, however due to the imbalance there are two RFP-associated mRNAs left to be translated and thus RFP would be expressed in this situation.<br />
<br />
<br />
Similar comparator circuit set-ups would be used in the ''Cura'' tattoo. For instance in the case of Type I Diabetes one promoter would sense blood sugar while another would sense insulin; if the blood sugar outweighed the insulin, more of the mRNA with the blood sugar promoter would be produced and thus an effector enzyme for the production of insulin could be included in this mRNA in order to increase insulin production. If insulin production becomes too high the imbalance would shift so more of the mRNA with the insulin-sensing promoter was produced, and an effector enzyme to break down excess insulin could be translated from this mRNA strand in order to normalise insulin levels. A system such as this, as well as others (which simply report changes through fluorescent proteins), are explored in Figure 2.<br />
<br />
<br />
[[File:NRPCura.png | center | 950 px | thumbnail | '''Figure 2.''' ''Cura'' is Quanticare's revolutionary healthcare solution, a tattoo made of immortalised mammalian cells containing different biobricks that have been specifically tailored to monitor and respond to changes in the body. Different areas of the tattoo correspond to different areas of the body, and using biological technology developed by the Norwich Research Park and University of East Anglia 2012 iGEM Team can react automatically to many unwanted changes, as well as fluorescing to alert the user of any problems.]]<br />
<br />
<br />
We have also looked at therapeutic strategies within Quanticare and how the tattoo may be analysed. As part of the film we looked at the idea of a scanner to complement the ''Cura'' tattoo, which will scan the tattoo's fluorescence patterns and feed through to a specialised printer designed to produce a bacterial pill optimised to treat the conditions reported by the tattoo. These bacteria would contain biobricks to produce drugs/enzymes as well as targeting mechanisms in order to find the site of a problem and correct mistakes being made; they could also be used to help swamp the likes of cancerous tumours and pump the tumour with drugs/apoptosis-inducing factors such as nitric oxide in order to help fight cancer; a mocked-up ''in vitro'' video of this occurring is included in the film. <br />
<br />
<br />
Through the film we have also explored the ethics of both synthetic biology healthcare solutions as well as synthetic biology as a whole. We have envisaged a world where public protests regarding synthetic biology have occurred, leading to companies such as Quanticare adopting stringent safety methods and testing in order to help calm public unease. We have also looked at the ethics of a tattoo and tattooing with immortalised cells, in particular the obvious risk of adding foreign cells to a body and public opinion on tattooing. We believe that while there is definite opposition, ''Cura'' is still a realistic future product provided the appropriate safety concerns are addressed (e.g. including a suicide gene in all cells should they begin to divide uncontrollably).<br />
<br />
<br />
As part of a marketing strategy to promote Quanticare and our film we have produced transferable tattoo versions of ''Cura'' which we will be handing out at the European Jamboree for other competitors, judges and interested parties to enjoy. We believe this will be an interactive and exciting approach to promoting our project and our ideas on the future of synthetic biology.</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/NOSensingTeam:NRP-UEA-Norwich/NOSensing2012-09-26T22:21:11Z<p>RussellGritton: /* Future Experiments */</p>
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<html><center><b><font size=4pt>Our hybrid promoter hopes to add to the systems already in the registry by creating a hybrid promoter that combines the bacterial promoter PyeaR and the mammalian CArG element , both of which respond to exogenous nitrogenous species. Combining the two would allow a more modular nitric oxide sensor that can be used in mammalian and bacterial cells interchangeably.</font> <br />
<br><br><br />
<font size=2pt>Six new biobricks produced and submitted to the registry with characterisation from fluorescence-based experiments.</font></b></center></html><br />
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Parts produced from this project:<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian/B-M (PyeaR-CArG) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial/M-B (CArG-PyeaR) Hybrid Promoter] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] -- [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP]<br />
<br />
Parts characterised from this project:<br />
<br />
[http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 PyeaR Promoter] -- [http://partsregistry.org/Part:BBa_K381001 PyeaR Promoter + GFP] -- [http://partsregistry.org/Part:BBa_E0420 enhanced Cyan Fluorescent Protein + RBS + Terminators] -- [http://partsregistry.org/Part:BBa_K081014 Red Fluorescent Protein + RBS + Terminators]<br />
<br />
Our main project has resulted in the production of a hybrid bacterial and mammalian promoter optimised for induction by nitric oxide, nitrates and nitrites. We have ligated PyeaR, a known bacterial promoter and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005] (Cambridge 2009) in the parts registry, with its mammalian counterpart, CArG. The resulting hybrid promoter has been synthesised in two orientations; PyeaR (bacterial, B) upstream of CArG (mammalian, M), nicknamed (B-M); and CArG upstream of PyeaR (M-B). These orientations were submitted to the parts registry as our first two biobricks.<br />
<br />
Each orientation of the promoter was ligated to enhanced cyan fluorescent protein (eCFP) and red fluorescent protein (RFP) to produce four new biobricks which have been submitted to the parts registry. These promoter + fluorescent protein biobricks have been characterised following transformation into ''Escherichia coli'' and induction by potassium nitrate using methods such as flow cytometry, fluorescence-activated cell sorting (FACS) and scanning with a fluorometer. The data from these experiments has proved that our promoter works as we expected it to. We have also transfected M-B + eCFP into a human breast cancer cell line, MCF7, and have proved the system is flexible and can be used in both eukaryotes and prokaryotes.<br />
<br />
We believe the promoters we have produced have relevant uses in cancer therapeutics, soil fertilisation and detection of emissions from industries such as construction.<br />
<br />
<br />
<br />
==Introduction==<br />
<br />
[[File:NRPPyeaR.png | 300px | thumbnail | '''''Figure 1.''''' ''A graphical representation of PyeaR. In the higher image PyeaR's activity is being repressed by both Nar and NsrR preventing transcription and the ultimate expression of a reporter. In the lower image nitrate/nitrite molecules have inhibited the activity of Nar, and nitric oxide has inhibited activity of NsrR, allowing for transcription to occur and subsequent expression of a reporter.'']]<br />
<br />
The University of East Anglia hosts many research teams whose work focuses on studying nitrogenous species and the way in which bacteria use and modify those species. One major problem that is faced by these teams involves working with nitric oxide (NO), as NO is highly reactive with a low half life, therefore making it difficult to trace and quantitatively measure accurately. Many of the methods currently used to measure NO levels are unable to distinguish between homogenous species, such as nitrates (NO3) and nitrites (NO2), therefore the figure given for NO levels is often inaccurate as other nitrogenous species are taken into account.<br />
<br />
<br />
The ability to be able to accurately detect NO levels is one with a great deal of potential for the future. Nitric oxide has been noted as a possible cancer therapy due to its physiological use as an apoptosis inducer by macrophages, however NO is also known to be used by cancerous cells to establish a baseline and use it to induce apoptosis and promote proliferation of a tumour; being able to accurately sense nitric oxide and go on to act on that information could be very useful to prevent the NO baseline being established by cancerous cells, but to also use NO for its apoptosis-inducing abilities. There are also other potential applications in the construction business, in 2008 Modern Building Services published an article regarding legislation released by the Building Regulation Establishment Environmental Assessment Method (BREEAM) encouraging construction companies to monitor their NO output as emission of NO can contribute to smog and acid rain levels; the ability to accurately detail levels of NO being released in these circumstances would be highly useful.<br />
<br />
<br />
Nitric oxide is an extremely physiologically relevant molecule found within both eukaryotes and prokaryotes, where it is utilised by different enzymes and systems for various roles. The aim of the experiment was to devise a hybrid promoter that could be applied to eukaryotes and prokaryotes in order to begin to more accurately sense NO and report on its specific levels.<br />
<br />
<br />
[[File:NRPBMandMB.png | 300px | thumbnail | '''''Figure 2.''''' ''A graphical representation of the hybrid promoter in its two orientations; the top image shows B-M with the bottom image showing M-B. Also included are the restriction sites and their location within the system, with the elements being read from left to right.'']]<br />
<br />
The hybrid promoter was designed to take on both bacterial and mammalian elements in order to be compatible with both bacterial and mammalian cells. After research around the subject and searching the parts registry a promoter known as PyeaR was decided upon as the bacterial element of the hybrid promoter; PyeaR is found in the ''yeaR/yoaG'' operon of ''Escherichia coli'' and is associated with induction by nitric oxide, nitrates and nitrites (Lin ''et al.'', 2007). PyeaR is repressed by two main repressors; Nar, which is regulated by nitrates and nitrites; and NsrR, which is regulated by nitric oxide (Figure 1.). One of the key elements of PyeaR is that it is not repressed in aerobic conditions, allowing for easier carrying out of experiments. The PyeaR aspect of the hybrid promoter has been known throughout the project as the bacterial promoter, or simply B.<br />
<br />
<br />
The mammalian element of the hybrid promoter was produced by nine CArG elements (repeated elements of CC(A/T)(6)GG), a promoter previously used synthetically for nitric oxide synthase as a cancer therapy (Worthington ''et al.'', 2005) and developed from the EGR1 gene for early growth response protein 1 (Scott ''et al.'', 2002). The CArG aspect of the hybrid promoter has been known throughout the project as the mammalian promoter, or simply M.<br />
<br />
<br />
Following identification of the two elements of the hybrid promoters the B (PyeaR) and M (CArG) aspects were ligated to one another in two orientations; B upstream of M (B-M) and M upstream of M (M-B) (Figure 2.). The hybrid promoters were synthesised in a pUC57 backbone with the standard iGEM restriction sites of EcoR1/Xba1 upstream of the promoter, and Spe1/Pst1 downstream of the promoter. A BamH1 restriction site was included in between the B and M sequences in order to allow for the B and M elements to be separated, as well as for easy verification of the promoter having been ligated into the iGEM backbone in future experiments (as BamH1 does not already exist in the pSB1C3 backbone).<br />
<br />
==Characterisation of Existing Biobrick: BBa_K381001 (PyeaR + GFP biobrick)==<br />
<br />
In order to begin to develop experiments to characterise the hybrid promoters + fluorescent proteins experiments were also carried out on a biobrick containing PyeaR + GFP (Part [http://partsregistry.org/Part:BBa_K381001 BBa_K381001], Bristol 2010). In these experiments transformed ''E. coli'' was inoculated into liquid culture, which in turn had varying potassium nitrate concentrations added to it. They were then left to grow before being spun down and viewed under a UV box in order to observe. The different concentrations of potassium nitrate that the transformed ''E. coli'' was grown in were: 0 mM, 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 8 mM, 10 mM.<br />
<br />
[[File:GFP 4.JPG | 600 px | center | thumbnail | '''''Figure 3.''''' ''A photograph of spun-down media containing potassium nitrate (to induce the promoter) and E. coli transformed by PyeaR + GFP (art BBa_K381001). Each sample was grown with a different concentration of potassium nitrate, from left to right: 0 mM, 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 8 mM, 10 mM.]]<br />
<br />
The figure suggests that fluorescent proteins have been expressed by the bacteria grown in media containing potassium nitrate due to the fluorescence shown under the UV box. It also suggests that different concentrations of potassium nitrate correlate with different intensities of expression due to the observable differences in fluorescence as the tubes are viewed from left to right, going up the gradient. The negative control of 0 mM potassium nitrate appears to show no fluorescence, suggesting it is indeed the potassium nitrate that is inducing the promoter.<br />
<br />
<br />
==Creating Novel Hybrid Promoters: B-M and M-B into Biobricks==<br />
<br />
The DNA for the synthesised genes of B-M and M-B had been supplied in the pUC57 backbone, therefore it was necessary for B-M and M-B to be digested from the pUC57 backbone and ligated into the pSB1C3 backbone. The synthesised gene was transformed into competent ’’E. coli’’ cells, which in turn were grown on agar plates containing 100 µg/ml ampicillin (due to pUC57 containing ampicillin resistance); colonies that had grown were then inoculated into liquid culture, and the liquid culture was subsequently mini-prepped using either the Bioline ISOLATE Plasmid DNA Mini Kit or the Promega Wizard® Plus SV Minipreps DNA Purification System. The DNA that had been extracted through mini-preps and the pSB1C3 backbone, as provided by the iGEM registry, were then digested using EcoR1 and Pst1 and a ligation was carried out using standard assembly protocol. The product of ligation was then transformed into competent ''E. coli'' cells, which were grown on agar plates containing 2.5 µg/ml chloramphenicol (due to pSB1C3 containing chloramphenicol resistance); this was done to eliminate any bacteria that had been transformed with undesirable ligation products. <br />
<br />
The colonies that had grown were then grown in liquid culture and mini-prepped in order to extract the DNA; the extracted DNA was then sent for sequencing, and the returned sequenced matched the expected sequence. The DNA was then sent to the parts registry as the [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774000 Bacterial-Mammalian Hybrid Promoter] and the [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774001 Mammalian-Bacterial Hybrid Promoter]. The biobricks for B-M and M-B were then used for further experiments, including ligation with a fluorescent protein reporter and growth studies.<br />
<br />
From weeks one through to five the team worked on producing the first biobricks of the hybrid promoter. Despite this proving difficult due to various reasons such as low amounts of DNA being produced from early mini-preps and ligation strategies not working, ''E. coli'' transformed by the biobrick DNA was successfully grown and proven to have the relevant antibiotic resistance by the beginning of [https://2012.igem.org/Team:NRP-UEA-Norwich/Week6 week six].<br />
<br />
===Studies into the effect of the hybrid promoter on growth of ''E. coli'' competent cells===<br />
<br />
[[File:NRPCalibration.png | 300 px | thumbnail | '''''Figure 4.''''' ''A calibration curve produced using E. coli alpha cells grown over time. The inoculations were run through a spectrophotometer at OD 600 nm every hour and then a sample was plated in order to calculate CFU/ml over time.]]<br />
<br />
As part of the characterisation of the hybrid promoters, as well as PyeaR alone ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K216005 Part BBa_K216005], studies were conducted to see if the growth of the ''E. coli'' was affected by transformation of the promoters. <br />
<br />
<br />
To being untransformed ''E. coli'' were inoculated into liquid culture and left to grow overnight at 37 ºC before being diluted down. The diluted liquid cultures were run through a spectrophotometer at OD 600 nm and their absorbance noted, before each culture was plated and once again left overnight at 37 ºC; the next morning the colonies on the plates were counted, allowing a calibration curve of absorbance at OD 600 nm against colony-forming units per ml (CFU/ml) to be set up (Figure 4.)<br />
<br />
<br />
Following establishment of the calibration curve liquid cultures were made up of ''E. coli'' that was:<br />
<br />
. Untransformed (otherwise referred to as alpha cells)<br />
<br />
. Transformed by B-M<br />
<br />
. Transformed by M-B<br />
<br />
. Transformed by PyeaR<br />
<br />
<br />
These cultures were then diluted down to the same starting level (an OD 600 nm absorbance of 0.2 +/- 0.1) and cuvettes filled with LB media were inoculated. The cuvettes were then placed in a spectrophotometer every hour and the absorbance at 600 nm was established; in between readings the cuvettes were placed into a 37 ºC incubator in order to encourage bacterial growth. This was repeated for 12 hours and the absorbance readings compared to the calibration curve in order to give data on the level of growth of ''E. coli'' transformed with the different promoters/untransformed over time.<br />
<br />
We found that there was a significant difference between Alpha cells and PyeaR cells. Initially, Alpha cells had a greater growth rate, but after the third hour into the study, the growth rate of PyeaR was faster than that of Alpha cells. The overall growth rate of PyeaR cells was significantly faster that Alpha cells (Levenes Test, F = 1.009 p = 0.372; T Test, t = 4.196, df = 4, p = 0.014).<br />
<br />
<br />
[[File:A + P.png | 600px | center | thumbnail | '''''Figure 5''''' ''Growth of PyeaR transformed E.coli cells relative to Alpha cell (untransformed cells. Error bars show the standard deviation between the three repeats. For clarity reasons, lines of best fit are not shown.'']]<br />
<br />
The growth pattern and rate of E.coli cells with or without transformation with B-M and M-B show little difference. Any differences in growth rate were not significant. There was lots of overlap. As previously described, there was a significant difference between the growth rate of PyeaR and Alpha cells. There was also a significant difference between MB/BM and PyeaR cells. The statistical results can be seen in Table 1<br />
<br />
[[File:A+M+B.png | 600px | center | thumbnail | '''''Figure 6:''''' ''Growth over 12 hours of Alpha, M-B and B-M. Error bars and lines of best fit are not shown for clarity reasons.'']]<br />
<br />
<br />
'''''Table 1:''''' ''ANOVA readings of statistical differences between Alpha (1) PyeaR (2), MB (3) and BM (4).''<br />
<br />
[[File:ANOVA.png | 600px | center]]<br />
<br />
From all the above graphs, it can be seen that with the starting concentration of cells as high as they are, the cultures are in exponential stage and do not undergo lag phase. A further growth study will be carried out on purely the lag phase with lower starting concentrations. As the starting absorbances here are approximately 0.2 at a wavelength of 600nm, the lag phase study will involve starting absorbances of 0.04 and lower.<br />
<br />
<br />
Following the above study, we found that a lag phase only study needed to be carried out to see if there was a significant difference in the lag phase. Again the study protocol was the same except that the starting concentration absorbances at 600nm was lowered to <0.04. It was extremely difficult to keep the absorbances ranges within 0.005 so the range is actually 0.3±0.1.<br />
<br />
[[File:PyeaR, BM, MB, alpha lag phase.png| 600px | center | thumbnail | '''''Figure 7''''' ''A mean average of all the data; using the data from the calibration curve, the absorbances were converted to colony forming units per ml (CFU/ml). The trend lines of alpha cells, BM/MB and PyeaR transformed cells are shown within this order from highest to lowest trendlines. One single trendline was used to represent BM and MB because the actual trendlines were extremely similar.'']]<br />
<br />
Using the initial concentrations of 0.3±0.1 showed that there is little difference between the growth rates. Using statistical analysis, it was found that there was no significant difference between any of the transformed cells relative to Alpha cells or to each other (Anova, p > 0.05). From this study we have found that changes in growth occur during exponential growth phase and not the lag growth phase.<br />
<br />
==Generation of B-M and M-B with eCFP and RFP==<br />
<br />
In order to test the activity of the hybrid promoters a reporter needed to be ligated. As the hybrid promoter did not already contain a ribosome binding site (RBS) both the RBS and the reporter were needed to be ligated to the promoter; in order to help improve experimental efficiency the parts registry was searched for relevant reporters that also contained an RBS. In [https://2012.igem.org/Team:NRP-UEA-Norwich/Week3 week three] two reporters were identified as [http://partsregistry.org/Part:BBa_E0420 BBa_E0420], a biobrick for enhanced CFP (eCFP) + RBS + terminators, and [http://partsregistry.org/Part:BBa_K081014 BBa_K081014], a biobrick for RFP + RBS + terminators.<br />
<br />
Once the B-M and M-B biobricks had been created in week six work began in earnest on the fluorescent proteins and ligating the promoters to them in order to begin characterisation. Due to many set-backs with low levels of DNA and having to order more biobricks from the registry, a successful ligation of the promoter to a fluorescent protein reporter was finally achieved in [https://2012.igem.org/Team:NRP-UEA-Norwich/Week10 week ten]. In order to carry out the ligation the promoter was first digested using Spe1 and Pst1 in order to linearise the backbone downstream of the promoter; the fluorescent proteins were digested using Xba1 and Pst1 in order to remove the insert. A ligation was then carried out using standard assembly protocol and the ligation products were transformed into ''E. coli'' competent cells, which in turn were grown on agar plates 2.5 µg/ml chloramphenicol (due to pSB1C3 containing chloramphenicol resistance). <br />
<br />
In order to quickly identify colonies of bacteria containing the promoter, RBS and reporter in a likely correct sequence a range of colonies were inoculated into media also containing potassium nitrate (KNO3) solution; this was done in order to inhibit the Nar repressor in PyeaR and result in activation of the promoter/induction of transcription/expression of the fluorescent protein reporter. Samples of the inoculated media containing KNO3 were then added to an eppendorf and spun down to form a pellet, which was viewed under a UV box and observed for fluorescence. After a week of various ligation and transformation experiments both promoters were successfully ligated to both fluorescent proteins and fluorescence of eCFP and RFP was observed under a UV box.<br />
<br />
From these experiments four more biobricks were produced and submitted to the registry: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 B-M + eCFP], [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP], [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP], and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774004 M-B + RFP].<br />
<br />
In order to characterise the hybrid promoters ligated to fluorescent proteins a number of experiments were carried out to measure the level of fluorescent output at different concentrations of KNO3 (used to induce the promoter's activity). All of these experiments were carried out in [https://2012.igem.org/Team:NRP-UEA-Norwich/Week11 week eleven]<br />
<br />
===Qualitative Results===<br />
<br />
[[File:NRPCFPWorks.jpg | 600 px | center | thumbnail | '''''Figure 8.''''' ''A photograph of spun-down media containing potassium nitrate (to induce the hybrid promoters) and E. coli transformed by M-B + eCFP. The concentrations of potassium nitrate added to the media were, from left to right: 100 mM, 50 mM, 10 mM, 0 mM.]]<br />
<br />
This figure appears to show fluorescence from the spun-down pellets of each hybrid promoter + fluorescent protein that had been grown in media contain potassium nitrate; it can be inferred from this that the potassium nitrate has induced the hybrid promoter, resulting in expression of the fluorescent protein reporter. The negative control of 0 mM potassium nitrate appears to show no fluorescence, suggesting it is indeed the potassium nitrate that is inducing the promoter.<br />
<br />
[[File:NRPFluorescence.jpeg | 600 px | center | thumbnail | '''''Figure 9.''''' ''A photograph of spun-down media containing potassium nitrate (to induce the hybrid promoters) and E. coli transformed by the four biobricks containg the promoters and fluorescent proteins; the photograph has been taken from a UV box. From left to right: B-M + eCFP, M-B + eCFP, B-M + RFP, M-B + RFP.]]<br />
<br />
The figure appears to show fluorescence from the spun-down pellets of each hybrid promoter + fluorescent protein that had been grown in media contain potassium nitrate; it can be inferred from this that the potassium nitrate has induced the hybrid promoter, resulting in expression of the fluorescent protein reporter.<br />
<br />
===Fluorometer Experiments===<br />
<br />
The main characterisation of the biobricks was carried out using a fluorometer. Five tubes of media containing 200 µL transformed bacteria and potassium nitrate were grown for each biobrick in concentrations as follows:<br />
<br />
. B-M + RFP with 0 mM, 5 mM, 10 mM, 15 mM and 20 mM potassium nitrate<br />
<br />
. B-M + eCFP with 0 mM, 5 mM, 10 mM, 15 mM and 20 mM potassium nitrate<br />
<br />
. M-B + RFP with 0 mM, 5 mM, 10 mM, 15 mM and 20 mM potassium nitrate<br />
<br />
. M-B + eCFP with 0 mM, 5 mM, 10 mM, 15 mM and 20 mM potassium nitrate<br />
<br />
<br />
The samples were then spun down in a centrifuge and resuspended in Tris buffer before being subjected to sonication in order to lyse the cells. The resulting solution was then spun down in a centrifuge again before the supernatant was extracted (in order to separate the proteins from the rest of the cell). The supernatant was then run through a fluorometer in order to measure the intensity of fluorescence as a method to gauge the activity of the hybrid promoters. RFP-containing samples were excited with a wavelength of 560 nm and the emission measured from 600 – 650 nm; eCFP-containing samples were excited with a wavelength of 410 nm and emission measured from 440-490 nm.<br />
<br />
<br />
These characterisation experiments gave rise to the characterisation which is now available on the main page for all hybrid promoter + fluorescent protein parts on the registry, and the experience section of the hybrid promoters alone on the registry.<br />
<br />
'''NOTE: All data for the fluorometer has had the equivalent 0 mM reading subtracted from it in order to nulify the affects of light scattering due to cell debris'''<br />
<br />
<br><br><br />
<br />
[[Image:BM-CFP_Graph.png | center | thumbnail | 600px | '''''Figure 10.''''' ''A graph of the intensity of CFP fluorescence at different wavelengths ranging from 440 - 500 nm where the samples were excited at 410 nm. The samples of E. coli were transformed by the B-M + CFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br>The graph above shows the flourescence measured from the expression of eCFP due to the response of the bacterial-mammalian promoter to different concentrations of potassium nitrate. The wavelength reading which corresponds to eCFP is between 440-500nm. The graph clearly demonstrates that between 0mN and 15mM there is a proportional relationship between fluorescence intensity and potassium nitrate concentration. There appears to be a sharp increase in fluorescence intensity between 5mM and 10mM, and the rate at which intensity increase gradually decreases so that there is only a small increase between 15mM and 20mM.<br />
[[Image:MB-CFP_Graph.png | center | thumbnail | 600px | '''''Figure 11.''''' ''A graph of the intensity of CFP fluorescence at different wavelengths ranging from 440 - 500 nm where the samples were excited at 410 nm. The samples of E. coli were transformed by the M-B + CFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br><br />
The graph above shows the flourescence measured from the expression of eCFP due to the response of the mammalian-bacterial promoter to different concentrations of potassium nitrate. The wavelength reading which corresponds to eCFP is between 440-500nm. The graph clearly demonstrates that between 0mN and 15mM there is a proportional relationship between fluorescence intensity and potassium nitrate concentration. It can be noted that at a 20mM concentration the intensity of fluorescence sharply decreases back down to the level of 5mM potassium nitate concentration. This may be due to the cell overexpressing eCFP up to the point at which the excess protein begins to form inclusion bodies which can no longer fluoresce; alternatively, this could be due the potassium nitrate concentration reaching the critical concentration at which it becomes toxic to the cell. This data differs to the readings taken from the bacterial-mammalian promoter ligated to eCFP, as well as the hybrid promoters to RFP, which may suggest there is a difference in the molecular mechanisms that these promoters function by; however at this point the change in intensity at 20mM is inconclusive and is an area which we would like to look into further. <br />
<br />
<br><br><br />
<br />
[[File:CFP_Comparison_Graph.png| center | thumbnail | 600px | '''''Figure 12.''''' ''A graph comparing the intensity of CFP fluorescence at 475 nm where the samples were excited at 410 nm. The samples of E. coli were transformed by the B-M + CFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br><br />
We were initially unsure of the effect that the orientation of the bacterial (pYEAR) and the mammalian (CaRG) genes would have in gene expression, therefore we synthesised two hybrid promoters in the orientation bacterial-mammalian and mammalian-bacterial. The graph above compares the intensity of fluorescence of the two hybrid promoters (BBa_K774004 and BBa_K774006) ligated to eCFP. There is a distinct difference between the intensity of fluorescence produced by the bacterial-mammalian promoter and the mammalian-promoter which is something that we would like to look into further. It is particularly interesting that at an intensity of 109a.u. the mammalian-bacterial promoter returns to the same level of intensity as the apparent maxiumum of the bacterial-mammalian promoter at 40a.u.<br />
<br />
<br><br><br />
We also ligated both of our hybrid promoters to Red Fluorescent Protein (RFP), and the results can be seen below.<br />
<br />
<br><br><br />
[[File:BM-RFP_Graph.png | center | thumbnail | 600px | '''''Figure 13.''''' ''A graph of the intensity of RFP fluorescence at different wavelengths ranging from 600 - 650 nm where the samples were excited at 560 nm. The samples of E. coli were transformed by the B-M + RFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br><br />
The graph above shows the flourescence measured from the expression of RFP due to the response of the bacterial-mammalian promoter to different concentrations of potassium nitrate. The wavelength reading which corresponds to RFP is between 600-650nm. The graph clearly demonstrates that between 0mN and 15mM there is a proportional relationship between fluorescence intensity and potassium nitrate concentration. A similar pattern can be seen here as for the mammalian- bacterial promoter with eCFP as at a 20mM concentration the intensity of fluorescence sharply decreases, however the intensity here decreases down to a level between 10mM and 15mM potassium nitate concentration. There is also only a small difference between 5mM and 10mM potassium nitrate, which differs to the pattern seen with the bacterial-mammalian promoter ligated to eCFP. As previously stated, this may be due to the cell overexpressing eCFP up to the point at which the excess protein begins to form inclusion bodies which can no longer fluoresce; alternatively, this could be due the potassium nitrate concentration reaching the critical concentration at which it becomes toxic to the cell. This data differs to the readings taken from the bacterial-mammalian ligated to eCFP, as well as the hybrid promoters to RFP, which may suggest there is a difference in the molecular mechanisms that these promoters function by; however at this point the change in intensity at 20mM is inconclusive and is an area which we would like to look into further.<br />
<br><br><br />
<br><br><br />
[[File:MB-RFP_Graph.png | center | thumbnail | 600px | '''''Figure 14.''''' ''A graph of the intensity of RFP fluorescence at different wavelengths ranging from 600 - 650 nm where the samples were excited at 560 nm. The samples of E. coli were transformed by the M-B + RFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br><br />
The graph above shows the flourescence measured from the expression of RFP due to the response of the mammalian-bacterial promoter to different concentrations of potassium nitrate. The wavelength reading which corresponds to RFP is between 600-650nm. The graph clearly demonstrates that between 0mN and 15mM there is a proportional relationship between fluorescence intensity and potassium nitrate concentration. It has been found that for all biobricks apart from the mammalian-bacterial promoter ligated to eCFP at a 20mM concentration the intensity of fluorescence sharply decreases. <br />
<br><br><br />
[[File:RFP_Comparison_Graph.png | center | thumbnail | 600px | '''''Figure 15.''''' ''A graph comparing the intensity of RFP fluorescence at different wavelengths ranging at 610 nm where the samples were excited at 560 nm. The samples of E. coli were transformed by the B-M + RFP biobrick and grown overnight in different concentrations of potassium nitrate (0 mM, 5 mM, 10 mM, 15 mM and 20 mM) before being lysed in order to release proteins from the cells for fluorometer analysis.'']]<br />
<br><br><br />
As previously stated, we were initially unsure of the effect that the orientation of the bacterial (pYEAR) and the mammalian (CaRG) genes would have in gene expression, therefore we synthesised two hybrid promoters in the orientation bacterial-mammalian and mammalian-bacterial. The graph above compares the intensity of fluorescence of the two hybrid promoters (BBa_K774007 and BBa_K774005) ligated to RFP. There appears to be no pattern if the difference between the intensities of these two promoters; however both promoters do show a decrease in intensity at 20mM potassium nitrate and decrease from a maximum intensity of 82a.u. (bacterial-mammalian) and 66a.u. to approximately 36a.u.<br />
<br />
===Flow Cytometry===<br />
<br />
Three tubes of media were inoculated with E. coli transformed by the [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774005 B-M + RFP] or [http://partsregistry.org/wiki/index.php?title=Part:BBa_K774006 M-B + eCFP] biobrick. Each tube then had potassium nitrate added to it at different concentrations; 0 mM, 1 mM and 10 mM respectively. The E. coli were grown over night and then spun down, fixed in 4% PFA and re-suspened in 500ul PBS. The samples were then analysed in an Acuri C6 or BD Aria II flow cytometer.<br />
<br />
[https://2012.igem.org/Team:NRP-UEA-Norwich/Protocol Full Protocol]<br />
<br />
''Note: This was the first time the flow cytometers at the University of East Anglia had been used with E. coli''<br />
<br />
====B-M + RFP Flow Cytometry Data====<br />
<br />
[[File:BM-RFP_18-9-12.png | 300 px | center | thumbnail | '''''Figure 16.''''' ''Flow cytometry data for B-M RFP transformed E. coli that were grown in either 0 mM, 1 mM or 10 mM potassium nitrate. Top row: Scatter plot of raw data and gating strategy utilised. Middle row: RFP Fluorescence profiles of samples. Lower left: Fluorescence profiles of the three samples overlay on the same plot.]]<br />
<br />
<br />
In this figure the image on the lower left of the fluorescence profiles overlayed on one another suggests that there is a slight difference in fluorescence intensity between the samples grown in media containing 0 mM potassium nitrate and 1 mM potassium nitrate, however there is a very noticeable difference in fluorescence between samples grown in 1 mM potassium nitrate and 10 mM potassium nitrate.<br />
<br />
[[File:BM-RFP.jpg | 300 px | center | thumbnail | '''''Figure 17.''''' ''Flow cytometry fluorescence data: B-M RFP transformed E. coli that were grown in either 0 mM, 1 mM or 10 mM potassium nitrate.]]<br />
<br />
The data in this figure appears to corroborate with the data in Figure 13, showing a small difference in fluorescence between 0 mM potassium nitrate and 1 mM potassium nitrate, and then a much larger difference in fluorescence intensity following induction by 10 mM potassium nitrate.<br />
<br />
<br><br />
<br />
====M-B + eCFP Flow Cytometry Data====<br />
<br />
[[File:MB-CFP_data.jpg | 300 px | center | thumbnail | '''''Figure 18.''''' ''Flow cytometry data for M-B eCFP transformed E. coli that were grown in either 0 mM, 1 mM or 10 mM potassium nitrate. Top row: Scatter plot of raw data and gating strategy utilised. Middle row: RFP Fluorescence profiles of samples.]]<br />
<br />
The data in this figure suggests a slight increase in fluorescence intensity from 0 mM to 1 mM, and then a higher increase in fluorescence intensity from 1 mM to 10 mM.<br />
<br />
[[File:MB-CFP.png | 300 px | center | thumbnail | '''''Figure 19.''''' ''Flow cytometry fluorescence data: M-B eCFP transformed E. coli that were grown in either 0 mM, 1 mM or 10 mM potassium nitrate.]]<br />
<br />
The data in this figure corroborates with the data in figure 15 in suggesting a slight increase in fluorescence intensity from 0 mM to 1 mM, and then a higher increase in fluorescence intensity from 1 mM to 10 mM.<br />
<br><br />
<br />
==Transfection of M-B + eCFP into the MCF7 human breast cancer cell line==<br />
<br />
In order to ascertain the flexibility of the hybrid promoter and help prove our original aim of producing a promoter that could be used in both eukaryotes and prokaryotes correct, the DNA for M-B + eCFP was transfected into the MCF7 human breast cancer cell line. An experiment was set up using S-Nitroso-N-acetyl-DL-penicillamine (SNAP), a nitric oxide donor, in order to induce expression of the fluorescent protein. A six-well transfection slide was produced containing:<br />
<br />
[[File:slide wells labelled.png | 500px | center]]<br />
<br />
The cells were then left for a day before they were imaged with a fluorescence microscope in order to observe expression of eCFP.<br />
<br />
[[File: Transfection.png | thumb | 500px |center | '''''Figure 20.''''' '' Transfection of MCF7 cells with images taken via a Zeiss CCD2 inverted microscope to detect CFP expression. Images in the left two columns are controls and have not been transfected, images in the right two columns have been transfected with M-B + CFP DNA; SNAP is a nitric oxide donor, therefore addition of SNAP was used to try and induce promoter activity.'']]<br />
<br />
The figure appears to show fluorescence in the mammalian cells that had been transfected with M-B + CFP compared with the cells that had not been transfected. The figure also appears to show stronger fluorescence in the cells that had been transfected with M-B + CFP and had been grown with the nitric oxide donor SNAP compared to the cells that had been transfected with M-B + CFP and grown without SNAP.<br />
<br />
[[File: NRPMBCFP.JPG | thumb | 500px |center | '''''Figure 21.''''' '' Transfection of MCF7 cells with images taken via a Zeiss CCD2 inverted microscope to detect CFP expression. Image shows mammalian cells transfected with M-B + CFP DNA however no SNAP (a nitric oxide donor) has been administered.'']]<br />
<br />
[[File:MCF7 blue.png | 300px | center ]]<br />
<br />
Close up image of what appears to be eCFP fluorescence from a transfected MCF7 cell.<br />
<br />
The figure appears to show low levels of fluorescence in the cells as there are small blue circles appearing in the cells indicative of CFP expression. MCF7 cells do endogenously express endothelial cell nitric-oxide synthase (ecNOS), which can produce low levels of NO. Perhaps it is endogenous NO production which is inducing the M-B promoter to express low levels of eCFP.<br />
<br><br />
<br />
[[File: NRPMBCFPSNAP.JPG | thumb | 500px |center | '''''Figure 22''''' '' Transfection of MCF7 cells with images taken via a Zeiss CCD2 inverted microscope to detect CFP expression. Image shows mammalian cells transfected with M-B + CFP DNA where SNAP (a nitric oxide donor) has been administered.'']]<br />
<br />
<br />
The figure suggests that expression of CFP has occurred due to the large blue shape appearing in the centre of the image following the use of a filter to view fluorescent cyan more easily.<br />
<br />
Future experiments are needed to confirm these results and also to provide more information.<br />
<br />
==Discussion==<br />
<br />
Through the experiments detailed above a lot of information has been collated regarding the hybrid promoters and their activity when induced with potassium nitrate. Through all figures from the fluorometer, flow cytometry and qualitative results showing fluorescence as a result of potassium nitrate induction we can conclude that the hybrid promoter in both orientations (B-M and M-B) does indeed react to nitrates and result in translation and expression of the downstream reporter. We can assume due to the transcriptional repressors associated with PyeaR (Figure 1.) that similar results would be given by nitrites, however future experimentation is recommended to test this theory and fully characterise this aspect of the promoter.<br />
<br />
<br />
In Figures 20, 21 and 22 it is shown that human breast cancer cells (MCF7) can be transfected with the M-B orientation of the promoter and that fluorescence does not occur in untransfected cells, however does occur in the transfected cells; the figures also appears to show that fluorescence intensifies upon addition of a nitric oxide donor. This data would suggest that the hybrid promoter indeed responds to nitric oxide as well as nitrates, however future experiments are needed to prove this. This appears to show that the hybrid promoter is indeed flexible as we first intended it to be, and systems can be developed in bacteria and then applied to mammalian cells with similar affects.<br />
<br />
<br />
In Figures 11, 13 and 14 the data shows the intensity of fluorescence increasing as the concentration of potassium nitrate added to the media the ''E. coli'' were grown in increases; this trend continues until the 15 mM sample (which appears to be the highest), before dramatically dropping in the case of the 20 mM potassium nitrate samples. One of the ideas surrounding this would be the toxicity levels of potassium nitrate, however in Figure 5 ''E. coli'' samples are shown to be fluorescing (and thus likely still be alive) after being subjected to 100 mM potassium nitrate. Another possibility may be the formation of inclusion bodies due to the vast quantities of fluorescent proteins being produced, leading to their aggregation and thus inability to fluoresce as expected. From the data given it can likely concluded that the maximum level of fluorescence can be attained from potassium nitrate levels between 15 mM and 20 mM, though future experimentation with narrower parameters in this area will be needed to conclude where the maximum tolerable level of potassium nitrate is found.<br />
<br />
<br />
In Figure 12 there appears to be a stark difference in CFP fluorescence intensity when M-B and B-M are compared, with M-B giving much higher intensities than B-M on all occasions, however in Figure 15 where the same experiments have been carried out using RFP instead of CFP the differences are not conclusively favouring M-B or B-M as the more active promoter. It is possible, therefore, that in the CFP experiments that there was a difference in the amount of cultured media that was used to inoculate fresh tubes (with more ''E. coli'' transformed by M-B + CFP being added than those transformed with B-M + CFP, resulting in more growth and thus increased production of CFP).<br />
<br />
<br />
In Figure 7 and through statistical analysis it is shown that there is no significant difference between the growth of cells transformed by M-B, B-M or PyeaR compared to untransformed cells; this shows that the hybrid promoter does not affect the growth of its chassis.<br />
<br />
<br />
'''Following the experiments over the 11-week project we have successfully produced two hybrid promoters including bacterial and mammalian elements in two orientations that sense nitrates and nitric oxide (and therefore likely nitrites). We have also successfully ligated the promoters to CFP and RFP as reporters, and have successfully transfected mammalian cells with M-B + CFP, proving the flexibility of the system.'''<br />
<br />
==Future Experiments==<br />
<br />
'''Full quantitative analysis to see where the values lie'''<br />
<br />
In order to gauge how the promoter responds to different levels of substrates experiments must be first designed to test the sensitivity to nitrates, nitrites and nitric oxide. The experiments detailed above have characterised a great deal of the nitrate sensitivity; moving on from this a salt such as sodium nitrite could be used as a donor for nitrites to assess how the promoter responds to nitrites as a substrate. A donor for nitric oxide must be found as well in order to ascertain more easily how the promoter responds to this compound (nitrite reductase has been looked at as a possibilty thus far). Finely-tuned experiments will be carried out to find the area at which fluorescence intensity moves from steadily increasing to dramatically dropping off (e.g. 15 mM - 20 mM for nitrates), and once a range has been found experiments would be carried out with small incriments of change of substrate concentration. Repeats will be made of all experiments in order to more accurately detail the change in intensity compared to substrate concentration. <br />
<br />
<br />
'''Combining the system with different reporter/effector enzymes'''<br />
<br />
Now we have completed experiments involving fluorescent protein reporters the next logical step for our future applications would be to attach different reporters or effector enzymes. We could attach an enzyme such as nitrite reductase in order to increase expression of nitric oxide as a next step experiment in the future application of cancer therapeutics; we could also investigate adding an enzyme to break down nitric oxide in order to take the next step in the future application of combating nitric oxide pollution. One idea would be to produce a new biobrick of M-B + Nitric Oxide Synthase (or another nitric oxide donor) + eCFP to test the concept of generating a synthetic gene network which can act as a cancer theraeutic.<br />
<br />
<br />
'''Repeat M-B + eCFP transfection into MCF7 cells. Utilise more experimental controls (e.g. M-B only transfection) and construct an M-B + eCFP plasmid which is optimised for mammalian systems'''<br />
<br />
We are very pleased with the results of our mammalian cell transfection, however it was just step one of what is likely very many more steps to optimise the experiment. In the future we would like to test transfection with M-B and B-M to see if both produce similar results in mammalian cells as well as using them as controls to promoters combined with reporters. We would also like to ligate the promoter into a plasmid which is more optimised for mammalian systems in order to test it fully within MCF7 cells.<br />
<br />
<br />
'''Optimise use of SNAP NO donor and use alternatives to enhance the induction of Nitric Oxide production'''<br />
<br />
In order to fully test nitric oxide sensation with the promoter we will need to test different nitric oxide donors e.g. SNAP, nitrite reductase, nitric oxide synthase. This will help us fully characterise how the promoter reacts the nitric oxide levels as well as help us decide on the best nitric oxide donor moving forward into producing systems for the future applications that will require nitric oxide production (e.g. cancer therapeutics)<br />
<br />
==References==<br />
<br />
Lin H.Y., Bledsoe P.J., Stewart V., (2007), ''Activation of yeaR-yoaG Operon Transcription by the Nitrate-Responsive Regulator NarL Is Independent of Oxygen- Responsive Regulator Fnr in Escherichia coli K-12▿'', Journal of Bacteriology, '''189: 7539 - 7548'''<br />
<br />
<br />
Modern Building Services, (2008), ''Understanding NOx emissions [online]'', Available at: [http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html], accessed on 26/09/2012<br />
<br />
<br />
Scott S.D., Joiner M.C., Marples B., (2002), ''Optimizing radiation-responsive gene promoters for radiogenetic cancer therapy.'', Gene Therapy, '''9: 1396-1402'''<br />
<br />
<br />
Worthington J., Robson T., Scott S., Hirst, D., (2005), ''Evaluation of a synthetic CArG promoter for nitric oxide synthase gene therapy of cancer'', Gene Therapy, '''12: 1417–1423'''</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/FutureTeam:NRP-UEA-Norwich/Future2012-09-26T21:43:13Z<p>RussellGritton: /* Understanding Pathogens */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
Nitric oxide (NO) is produced in different environments, be that within humans or by bacteria in soil. It is a nitrogenous species that is involved in the nitrogen cycle. With further research, development and integration into various systems, the future of medicine and agriculture could involve NO sensors.<br />
<br />
=='''NO sensing'''==<br />
<br />
Many NO sensors exist, but usually have problems with specific and quantitative NO detection (Bedioui, F .et al.,2002). To do this in a mlore specific manner, a comparator circuit system can be put into place. The circuit could involve multiple sensors that sense different reactive oxygen and nitrogenous species. Within this system, short interfering RNA (siRNA) could be implemented to inhibit the expression of certain enzymes and proteins. This would produce an overall output that corresponds only to NO levels. To give a visual quantitative output, the use of bioluminescence can be augmented. The brightness would relate to the amount of NO in cells environment.<br />
<br />
Furthermore, as mentioned previously, the E.chromi project can be incorporated. In addition to its application in disease communication relative to NO levels, different sensors can be incorporated into the system to accurately measure all nitrogenous compounds. The levels of these can be qualitatively measured through the resulting coloured product.<br />
<br />
[[File:Future applications.png|950px]]<br />
<br />
=='''Agriculture'''==<br />
<br />
NO can be produced by nitrifying bacteria during oxidation of ammonium (Lipschultz, F. et al. 1981). These gases can be released into the atmosphere. NO is also converted back to atmospheric nitrogen in a reductive process called denitrification. The differences in the bacterial species that compose soil flora produce and reduce nitric oxide lead to different levels of NO in soil, which can affect soil fertility and thus agricultural yield. Besides soil flora, soil composition, temperature, water levels (Davidson, E. 2012), tilling and fertiliser quantities (Civerolo, K.L. et al. 1998) can all affect the levels of NO. This is particularly significant now as the world population is ever increasing; the higher the agricultural yield the more people that can be supported. An accurate NO sensor can allow a farmer to be able to better utilise their resources and to accurately apply fertiliser, water and needed farming techniques to maximise the crop yield.<br />
<br />
=='''Medicine'''==<br />
<br />
[[File:JAMES_CAVALLINI_SCIENCE_PHOTO_LIBRARY.jpg| 200px | right | thumbnail | <html><u>This photograph is the property of James Cavallini/ Science Photo Library.</u></html>]]<br />
<br />
<br />
<br />
Nitric oxide is an important physiological signalling molecule within the human body. This highly reactive molecule is produced through the action of enzymes called nitic oxide synthases (NOS). There are three main types of NOS in the body, which each produce NO for a different physiological role: inducible (iNOS), endothelial (eNOS) and neuronal (nNOS) (Xu, W. et al. 2002). NO in the blood, causes vasodilation through increase of cGMP, a second messenger which activates many receptors and processes (Ferreira, L.F. et al 2010). As NO signalling is so widespread within the body there are many applications NO sensors can have in medicine.<br />
<br><br>Whilst NO triggers several defence mechanisms within the innate immune response, it has been found that some bacteria, such as the food-borne pathogen ''Listeria monocytogenes'', are able to use increased NO levels (produced by nitric oxide synthase-2) to promote their colonisation of host cells; culminating in a condition known as listeriosis. ''L. monoctogenes'' is absorbed from donor to recipient cell within vacuoles, and as NO decreases the rate at which the immune response destroys these vacuoles the bacteria is increasingly successful (Cole, C. et al. 2012). A system which is able to both detect and decrease levels of nitric oxide would thus be highly advantageous in therapeutic strategies to treat ''L. monocytogenes'' infection.<br />
<br />
<Br><Br><br />
<br />
<br />
=='''Cancer Research'''==<br />
<br />
[[File:Warrior_cell.png| 200px| left | thumbnail |This image is a logo which represents a future application of how our biobricks could be used to produce a bacteria capable of acting as a cancer thereapeutic.]]<br />
All the isoforms of NO synthases aforementioned are involved in the promotion or inhibition of the tumour cells (Xu, W. et al. 2002). Research has shown that high levels of NO can be cytotoxic /or cytostatic to tumour cells; however, at the low levels produced by tumour cells, NO signals angiogenic factors such as VEGF leading to angiogenesis and enlargement of the tumour (Xu, W. et al. 2002). Macrophages, as part of the innate immune system, produce NO to kill off the tumour. Research by Xu et al., shows that NO has greater activity when the tumour is less<br />
differentiated and hence a higher grade.<br />
<br />
Using NO as a chemoattractant, the low and high levels of NO produced by cancer cells and macrophages, respectively can be perceived by the sensor which can target drugs directly to the tumour. An alternative method is to use oxygen as a chemorepellent so modified cells can specifically target hypoxic cells. Specificity can be increased through the use of targeting using specific peptide sequences which home onto receptors or glycoproteins which are specific to tumour cells such as the NGR peptide (Pasqualini, R. et al. 2000).<br />
<br />
<br><br><br><br><br><br><br><br><br><br />
<br />
=='''Diagnostics'''==<br />
<br />
[[File:E.chromi.jpg| 200px| left | thumbnail |This image is from the Cambridge E.chromi project, showing the coloured faecal matter as a form of diagnostic which has potential for identifying damage to the gut.]]NO is produced throughout the mammalian body in a range of cells, including endothelial cells (Bedioi.F et,al., 2002). NO has a range of functions within the body, from being a secondary messenger in signalling pathways to being a relaxing factor within the cardiovascular system (Pang, et,al.,2003). Abnormal levels of NO can be an indication of diseases and disorders such as hypertension, impotence and obesity (Bedioui.F et,al.,2002). A NO sensor can be augmented into two ways to aid diagnosis and treatment. For example, measuring the NO level of externally exhaled air can be an important and non-invasive diagnostic for airways inflammation. This is due to endothelial cells producing NO synthase (eNOS), both constitutive and inducible (iNOS). The iNOS is induced by pro inflammatory cytokines, as well as there being an increase in enzyme expression after exposure to oxidants. Therefore, measuring the NO level produced by eNOS has led to finding that there are higher levels of epithelial nitric oxide when asthma is untreated, and that there is a fall after anti-inflammatory treatment (Narang ,et al., 2002).<br />
<br />
Alternatively,Building upon the Cambridge 2009 iGEM E.Chromi project, the NO sensor can be incorporated to perceive abnormal levels of NO in the gut flora or damage to the gut, which can visually be seen in the form of colours in faecal matter (Cambridge iGEM team , 2009). For example NO levels increase during damage of the gut, potentially due to its role in functional repair ( Miller et al, 1993). Therefore,an NO sensor would produce particular colours that could correspond to different levels of NO expected for certain diseases and disorders.<br />
<br />
Another use is through the cells being stimulated to create a protein coat for resistance and persistence within the human body. These cells could be mass produced and contain vaccines which can be taken at birth. These cells can be used to target pathogens or other abnormalities that arise in the body.<br />
<br />
=='''Understanding Pathogens'''==<br />
<br />
[[File:Sensing.jpg| 200px| left | thumbnail |(Henares, et, al., 2012). The process used by bacteria in which NO course quorum sensing through lux U within V.harveyi.]]<br />
Pathogens have many different mechanisms and strategies of invasion and attack. For example some pathogens produce biofilms. To create biofilms, bacteria signal to other bacteria to assess the population size and density (Tsou, et al., 2009). When numbers are sufficient, they switch on gene expression which leads to aggregation and adhesion. This is an example of quorum sensing, as has been demonstrated through Nitric Oxide responsive quorum sensing circuits though lux U within Vibrio harveyi(Henares, et al., 2012). A greater understanding of NO can be applied to research into pathogenic diseases which could lead to advances in the creation of cures and vaccines.<br />
<br />
<br><br><br />
<br />
=='''Construction and Buildings Emissions'''==<br />
<br />
In 2008 the Modern Buildings Services released an article detailing recommendations from the Buildings Regulation Establishment Environmental Assessment Method (BREEAM) encouraging construction and buildings companies to monitor levels of nitric oxide emissions. They detail nitric oxide formation through combustion reactions involved in certain construction methods and even household applicances (e.g. boilers) and the consequences of excess atmospheric nitric oxide, including the production of smog and acid rain. Production of a way to accurately sense nitric oxide levels in the form of the hybrid promoter would be an extremely useful analytical tool; further to this combining the hybrid promoter and the comparator circuit could see bacteria that could not only sense nitric oxide being released by these appliances etc., but could also express an enzyme to safely break nitric oxide down and prevent the formation of harmful compounds.<br />
<br><br><br />
<br />
==References==<br />
<br />
Bedioui,F., Villeneuue, N.(2002)‘ Electrochemical Nitric Oxide Sensors for biological samples- Principle, selected examples and applications’, Electroanalysis, 15; 5-18. <br />
<br><br><br />
Cambridge iGEM team (2009) Available at: https://2009.igem.org/Team:Cambridge [Accessed on: 21/09.2012] <br />
<br><br><br />
Civerolo, K.L. and Dickerson, R.R. (1998) ‘Nitric oxide soil emissions from tilled and untilled corn®elds’, Agricultural and Forest Meteorology, 90; 307-311. <br />
<br><br><br />
Cole, C., Thomas, S., Filak, H., Henson, P., Lenz, L. (2012) ‘Nitric Oxide Increases Susceptibility of Toll-like Receptor-Activated Macrophages to Spreading Listeria monocytogenes’, Immunity, 36; 885.<br />
<br><br><br />
Davidson, E. (2012) Sources of Nitric Oxide and Nitrous Oxide following Wetting of Dry Soil, Soil Sci. Soc. Am. J. 56; 95–102<br />
<br><br><br />
Ferreira, L.F. and Behnke, B.J. (2010) ‘A toast to health and performance! Beetroot juice lowers blood pressure and the O2 cost of exercise’, Journal of Applied Physiology, 110; 585-586.<br />
<br><br><br />
Henares, B.M., Higgins, K.E., Boon, E.M. (2012)'Discovery of a Nitric Oxide Responsive Quorum Sensing Circuit in Vibrio harveyi',ACS chemical biology,17;1331-6.<br />
<br><br><br />
Lipschultz, F., Zafiriou, O.C. Wofsy, S.C., Elroy, M.B., Valois, F.W. and Watson, S.W. (1981) ‘Production of NO and N2O by soil nitrifying bacteria’, Macmillan Journals, 294; 641-643.<br />
<br><br><br />
Miller, M.J., Zhang, X.J., Sadowska-Krowicka, H., Chotinaruemol ,S., McIntyre ,J.A., Clark ,D.A. and Bustamante ,S.A. (1993) 'Nitric oxide release in response to gut injury',scandinavian journal of gastroenterology,28; 149-54.<br />
<br><br><br />
Modern Building Services, (2008), ''Understanding NOx emissions [online]'', Available at: [http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html], accessed on 26/09/2012<br />
<br><br><br />
Narang, I., Ersu,R., Wilson, N.M., Bush. A. (2002) 'Nitric oxide in chronic airway inflammation in children: diagnostic use and pathophysiological significance', Thorax ,57;586-589.<br />
<br><br><br />
Pang.J, Fan.C, Liv.X, Chem.T, Li.G (2003)'A nitric oxide biosensor or based on the multi-assembly of hemoglobin, montmorillonite/ polyvinyl alcohol at pyrolytic graphite electrode.', Bio sensors and bioelectronics, 19: 441-445. <br />
<br><br><br />
Pasqualini, R., Koivunen, E., Kain, R., Lahdenranta, J., Sakamoto, M., Stryhn, A., Ashmun, R.A., Shapiro, L.H., Arap, W. And Ruoslahti, E. (2000) ‘Aminopeptidase N is a receptor for tumour-homing peptides and a target for inhibiting angiogenesis’, The Journal of Cancer Research, 60; 722-727.<br />
<br><br><br />
Tsou,A.M., Cai,T., Liu,Z.,Zhu,J., and Kulkarni, R.V.,(2009)'Regulatory targets of quorum sensing in Vibrio cholerae: evidence for two distinct HapR-binding motifs',Nucleic Acids Res, 37; 2747–2756.<br />
<br><br><br />
Xu, W., Liu, L.Z., Loizidou, M., Ahmed, M. And Charles, I.G. (2002) ‘The role of nitric oxide in cancer’, Cell Research, 12; 311-320</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/FutureTeam:NRP-UEA-Norwich/Future2012-09-26T21:42:41Z<p>RussellGritton: /* Construction and Buildings Emissions */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
Nitric oxide (NO) is produced in different environments, be that within humans or by bacteria in soil. It is a nitrogenous species that is involved in the nitrogen cycle. With further research, development and integration into various systems, the future of medicine and agriculture could involve NO sensors.<br />
<br />
=='''NO sensing'''==<br />
<br />
Many NO sensors exist, but usually have problems with specific and quantitative NO detection (Bedioui, F .et al.,2002). To do this in a mlore specific manner, a comparator circuit system can be put into place. The circuit could involve multiple sensors that sense different reactive oxygen and nitrogenous species. Within this system, short interfering RNA (siRNA) could be implemented to inhibit the expression of certain enzymes and proteins. This would produce an overall output that corresponds only to NO levels. To give a visual quantitative output, the use of bioluminescence can be augmented. The brightness would relate to the amount of NO in cells environment.<br />
<br />
Furthermore, as mentioned previously, the E.chromi project can be incorporated. In addition to its application in disease communication relative to NO levels, different sensors can be incorporated into the system to accurately measure all nitrogenous compounds. The levels of these can be qualitatively measured through the resulting coloured product.<br />
<br />
[[File:Future applications.png|950px]]<br />
<br />
=='''Agriculture'''==<br />
<br />
NO can be produced by nitrifying bacteria during oxidation of ammonium (Lipschultz, F. et al. 1981). These gases can be released into the atmosphere. NO is also converted back to atmospheric nitrogen in a reductive process called denitrification. The differences in the bacterial species that compose soil flora produce and reduce nitric oxide lead to different levels of NO in soil, which can affect soil fertility and thus agricultural yield. Besides soil flora, soil composition, temperature, water levels (Davidson, E. 2012), tilling and fertiliser quantities (Civerolo, K.L. et al. 1998) can all affect the levels of NO. This is particularly significant now as the world population is ever increasing; the higher the agricultural yield the more people that can be supported. An accurate NO sensor can allow a farmer to be able to better utilise their resources and to accurately apply fertiliser, water and needed farming techniques to maximise the crop yield.<br />
<br />
=='''Medicine'''==<br />
<br />
[[File:JAMES_CAVALLINI_SCIENCE_PHOTO_LIBRARY.jpg| 200px | right | thumbnail | <html><u>This photograph is the property of James Cavallini/ Science Photo Library.</u></html>]]<br />
<br />
<br />
<br />
Nitric oxide is an important physiological signalling molecule within the human body. This highly reactive molecule is produced through the action of enzymes called nitic oxide synthases (NOS). There are three main types of NOS in the body, which each produce NO for a different physiological role: inducible (iNOS), endothelial (eNOS) and neuronal (nNOS) (Xu, W. et al. 2002). NO in the blood, causes vasodilation through increase of cGMP, a second messenger which activates many receptors and processes (Ferreira, L.F. et al 2010). As NO signalling is so widespread within the body there are many applications NO sensors can have in medicine.<br />
<br><br>Whilst NO triggers several defence mechanisms within the innate immune response, it has been found that some bacteria, such as the food-borne pathogen ''Listeria monocytogenes'', are able to use increased NO levels (produced by nitric oxide synthase-2) to promote their colonisation of host cells; culminating in a condition known as listeriosis. ''L. monoctogenes'' is absorbed from donor to recipient cell within vacuoles, and as NO decreases the rate at which the immune response destroys these vacuoles the bacteria is increasingly successful (Cole, C. et al. 2012). A system which is able to both detect and decrease levels of nitric oxide would thus be highly advantageous in therapeutic strategies to treat ''L. monocytogenes'' infection.<br />
<br />
<Br><Br><br />
<br />
<br />
=='''Cancer Research'''==<br />
<br />
[[File:Warrior_cell.png| 200px| left | thumbnail |This image is a logo which represents a future application of how our biobricks could be used to produce a bacteria capable of acting as a cancer thereapeutic.]]<br />
All the isoforms of NO synthases aforementioned are involved in the promotion or inhibition of the tumour cells (Xu, W. et al. 2002). Research has shown that high levels of NO can be cytotoxic /or cytostatic to tumour cells; however, at the low levels produced by tumour cells, NO signals angiogenic factors such as VEGF leading to angiogenesis and enlargement of the tumour (Xu, W. et al. 2002). Macrophages, as part of the innate immune system, produce NO to kill off the tumour. Research by Xu et al., shows that NO has greater activity when the tumour is less<br />
differentiated and hence a higher grade.<br />
<br />
Using NO as a chemoattractant, the low and high levels of NO produced by cancer cells and macrophages, respectively can be perceived by the sensor which can target drugs directly to the tumour. An alternative method is to use oxygen as a chemorepellent so modified cells can specifically target hypoxic cells. Specificity can be increased through the use of targeting using specific peptide sequences which home onto receptors or glycoproteins which are specific to tumour cells such as the NGR peptide (Pasqualini, R. et al. 2000).<br />
<br />
<br><br><br><br><br><br><br><br><br><br />
<br />
=='''Diagnostics'''==<br />
<br />
[[File:E.chromi.jpg| 200px| left | thumbnail |This image is from the Cambridge E.chromi project, showing the coloured faecal matter as a form of diagnostic which has potential for identifying damage to the gut.]]NO is produced throughout the mammalian body in a range of cells, including endothelial cells (Bedioi.F et,al., 2002). NO has a range of functions within the body, from being a secondary messenger in signalling pathways to being a relaxing factor within the cardiovascular system (Pang, et,al.,2003). Abnormal levels of NO can be an indication of diseases and disorders such as hypertension, impotence and obesity (Bedioui.F et,al.,2002). A NO sensor can be augmented into two ways to aid diagnosis and treatment. For example, measuring the NO level of externally exhaled air can be an important and non-invasive diagnostic for airways inflammation. This is due to endothelial cells producing NO synthase (eNOS), both constitutive and inducible (iNOS). The iNOS is induced by pro inflammatory cytokines, as well as there being an increase in enzyme expression after exposure to oxidants. Therefore, measuring the NO level produced by eNOS has led to finding that there are higher levels of epithelial nitric oxide when asthma is untreated, and that there is a fall after anti-inflammatory treatment (Narang ,et al., 2002).<br />
<br />
Alternatively,Building upon the Cambridge 2009 iGEM E.Chromi project, the NO sensor can be incorporated to perceive abnormal levels of NO in the gut flora or damage to the gut, which can visually be seen in the form of colours in faecal matter (Cambridge iGEM team , 2009). For example NO levels increase during damage of the gut, potentially due to its role in functional repair ( Miller et al, 1993). Therefore,an NO sensor would produce particular colours that could correspond to different levels of NO expected for certain diseases and disorders.<br />
<br />
Another use is through the cells being stimulated to create a protein coat for resistance and persistence within the human body. These cells could be mass produced and contain vaccines which can be taken at birth. These cells can be used to target pathogens or other abnormalities that arise in the body.<br />
<br />
=='''Understanding Pathogens'''==<br />
<br />
[[File:Sensing.jpg| 200px| left | thumbnail |(Henares, et, al., 2012). The process used by bacteria in which NO course quorum sensing through lux U within V.harveyi.]]<br />
Pathogens have many different mechanisms and strategies of invasion and attack. For example some pathogens produce biofilms. To create biofilms, bacteria signal to other bacteria to assess the population size and density (Tsou, et al., 2009). When numbers are sufficient, they switch on gene expression which leads to aggregation and adhesion. This is an example of quorum sensing, as has been demonstrated through Nitric Oxide responsive quorum sensing circuits though lux U within Vibrio harveyi(Henares, et al., 2012). A greater understanding of NO can be applied to research into pathogenic diseases which could lead to advances in the creation of cures and vaccines.<br />
<br />
<br><br><br><br />
<br />
=='''Construction and Buildings Emissions'''==<br />
<br />
In 2008 the Modern Buildings Services released an article detailing recommendations from the Buildings Regulation Establishment Environmental Assessment Method (BREEAM) encouraging construction and buildings companies to monitor levels of nitric oxide emissions. They detail nitric oxide formation through combustion reactions involved in certain construction methods and even household applicances (e.g. boilers) and the consequences of excess atmospheric nitric oxide, including the production of smog and acid rain. Production of a way to accurately sense nitric oxide levels in the form of the hybrid promoter would be an extremely useful analytical tool; further to this combining the hybrid promoter and the comparator circuit could see bacteria that could not only sense nitric oxide being released by these appliances etc., but could also express an enzyme to safely break nitric oxide down and prevent the formation of harmful compounds.<br />
<br><br><br />
<br />
==References==<br />
<br />
Bedioui,F., Villeneuue, N.(2002)‘ Electrochemical Nitric Oxide Sensors for biological samples- Principle, selected examples and applications’, Electroanalysis, 15; 5-18. <br />
<br><br><br />
Cambridge iGEM team (2009) Available at: https://2009.igem.org/Team:Cambridge [Accessed on: 21/09.2012] <br />
<br><br><br />
Civerolo, K.L. and Dickerson, R.R. (1998) ‘Nitric oxide soil emissions from tilled and untilled corn®elds’, Agricultural and Forest Meteorology, 90; 307-311. <br />
<br><br><br />
Cole, C., Thomas, S., Filak, H., Henson, P., Lenz, L. (2012) ‘Nitric Oxide Increases Susceptibility of Toll-like Receptor-Activated Macrophages to Spreading Listeria monocytogenes’, Immunity, 36; 885.<br />
<br><br><br />
Davidson, E. (2012) Sources of Nitric Oxide and Nitrous Oxide following Wetting of Dry Soil, Soil Sci. Soc. Am. J. 56; 95–102<br />
<br><br><br />
Ferreira, L.F. and Behnke, B.J. (2010) ‘A toast to health and performance! Beetroot juice lowers blood pressure and the O2 cost of exercise’, Journal of Applied Physiology, 110; 585-586.<br />
<br><br><br />
Henares, B.M., Higgins, K.E., Boon, E.M. (2012)'Discovery of a Nitric Oxide Responsive Quorum Sensing Circuit in Vibrio harveyi',ACS chemical biology,17;1331-6.<br />
<br><br><br />
Lipschultz, F., Zafiriou, O.C. Wofsy, S.C., Elroy, M.B., Valois, F.W. and Watson, S.W. (1981) ‘Production of NO and N2O by soil nitrifying bacteria’, Macmillan Journals, 294; 641-643.<br />
<br><br><br />
Miller, M.J., Zhang, X.J., Sadowska-Krowicka, H., Chotinaruemol ,S., McIntyre ,J.A., Clark ,D.A. and Bustamante ,S.A. (1993) 'Nitric oxide release in response to gut injury',scandinavian journal of gastroenterology,28; 149-54.<br />
<br><br><br />
Modern Building Services, (2008), ''Understanding NOx emissions [online]'', Available at: [http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html], accessed on 26/09/2012<br />
<br><br><br />
Narang, I., Ersu,R., Wilson, N.M., Bush. A. (2002) 'Nitric oxide in chronic airway inflammation in children: diagnostic use and pathophysiological significance', Thorax ,57;586-589.<br />
<br><br><br />
Pang.J, Fan.C, Liv.X, Chem.T, Li.G (2003)'A nitric oxide biosensor or based on the multi-assembly of hemoglobin, montmorillonite/ polyvinyl alcohol at pyrolytic graphite electrode.', Bio sensors and bioelectronics, 19: 441-445. <br />
<br><br><br />
Pasqualini, R., Koivunen, E., Kain, R., Lahdenranta, J., Sakamoto, M., Stryhn, A., Ashmun, R.A., Shapiro, L.H., Arap, W. And Ruoslahti, E. (2000) ‘Aminopeptidase N is a receptor for tumour-homing peptides and a target for inhibiting angiogenesis’, The Journal of Cancer Research, 60; 722-727.<br />
<br><br><br />
Tsou,A.M., Cai,T., Liu,Z.,Zhu,J., and Kulkarni, R.V.,(2009)'Regulatory targets of quorum sensing in Vibrio cholerae: evidence for two distinct HapR-binding motifs',Nucleic Acids Res, 37; 2747–2756.<br />
<br><br><br />
Xu, W., Liu, L.Z., Loizidou, M., Ahmed, M. And Charles, I.G. (2002) ‘The role of nitric oxide in cancer’, Cell Research, 12; 311-320</div>RussellGrittonhttp://2012.igem.org/Team:NRP-UEA-Norwich/FutureTeam:NRP-UEA-Norwich/Future2012-09-26T21:42:14Z<p>RussellGritton: /* Understanding Pathogens */</p>
<hr />
<div>{{UEANRP}}<br />
<br />
Nitric oxide (NO) is produced in different environments, be that within humans or by bacteria in soil. It is a nitrogenous species that is involved in the nitrogen cycle. With further research, development and integration into various systems, the future of medicine and agriculture could involve NO sensors.<br />
<br />
=='''NO sensing'''==<br />
<br />
Many NO sensors exist, but usually have problems with specific and quantitative NO detection (Bedioui, F .et al.,2002). To do this in a mlore specific manner, a comparator circuit system can be put into place. The circuit could involve multiple sensors that sense different reactive oxygen and nitrogenous species. Within this system, short interfering RNA (siRNA) could be implemented to inhibit the expression of certain enzymes and proteins. This would produce an overall output that corresponds only to NO levels. To give a visual quantitative output, the use of bioluminescence can be augmented. The brightness would relate to the amount of NO in cells environment.<br />
<br />
Furthermore, as mentioned previously, the E.chromi project can be incorporated. In addition to its application in disease communication relative to NO levels, different sensors can be incorporated into the system to accurately measure all nitrogenous compounds. The levels of these can be qualitatively measured through the resulting coloured product.<br />
<br />
[[File:Future applications.png|950px]]<br />
<br />
=='''Agriculture'''==<br />
<br />
NO can be produced by nitrifying bacteria during oxidation of ammonium (Lipschultz, F. et al. 1981). These gases can be released into the atmosphere. NO is also converted back to atmospheric nitrogen in a reductive process called denitrification. The differences in the bacterial species that compose soil flora produce and reduce nitric oxide lead to different levels of NO in soil, which can affect soil fertility and thus agricultural yield. Besides soil flora, soil composition, temperature, water levels (Davidson, E. 2012), tilling and fertiliser quantities (Civerolo, K.L. et al. 1998) can all affect the levels of NO. This is particularly significant now as the world population is ever increasing; the higher the agricultural yield the more people that can be supported. An accurate NO sensor can allow a farmer to be able to better utilise their resources and to accurately apply fertiliser, water and needed farming techniques to maximise the crop yield.<br />
<br />
=='''Medicine'''==<br />
<br />
[[File:JAMES_CAVALLINI_SCIENCE_PHOTO_LIBRARY.jpg| 200px | right | thumbnail | <html><u>This photograph is the property of James Cavallini/ Science Photo Library.</u></html>]]<br />
<br />
<br />
<br />
Nitric oxide is an important physiological signalling molecule within the human body. This highly reactive molecule is produced through the action of enzymes called nitic oxide synthases (NOS). There are three main types of NOS in the body, which each produce NO for a different physiological role: inducible (iNOS), endothelial (eNOS) and neuronal (nNOS) (Xu, W. et al. 2002). NO in the blood, causes vasodilation through increase of cGMP, a second messenger which activates many receptors and processes (Ferreira, L.F. et al 2010). As NO signalling is so widespread within the body there are many applications NO sensors can have in medicine.<br />
<br><br>Whilst NO triggers several defence mechanisms within the innate immune response, it has been found that some bacteria, such as the food-borne pathogen ''Listeria monocytogenes'', are able to use increased NO levels (produced by nitric oxide synthase-2) to promote their colonisation of host cells; culminating in a condition known as listeriosis. ''L. monoctogenes'' is absorbed from donor to recipient cell within vacuoles, and as NO decreases the rate at which the immune response destroys these vacuoles the bacteria is increasingly successful (Cole, C. et al. 2012). A system which is able to both detect and decrease levels of nitric oxide would thus be highly advantageous in therapeutic strategies to treat ''L. monocytogenes'' infection.<br />
<br />
<Br><Br><br />
<br />
<br />
=='''Cancer Research'''==<br />
<br />
[[File:Warrior_cell.png| 200px| left | thumbnail |This image is a logo which represents a future application of how our biobricks could be used to produce a bacteria capable of acting as a cancer thereapeutic.]]<br />
All the isoforms of NO synthases aforementioned are involved in the promotion or inhibition of the tumour cells (Xu, W. et al. 2002). Research has shown that high levels of NO can be cytotoxic /or cytostatic to tumour cells; however, at the low levels produced by tumour cells, NO signals angiogenic factors such as VEGF leading to angiogenesis and enlargement of the tumour (Xu, W. et al. 2002). Macrophages, as part of the innate immune system, produce NO to kill off the tumour. Research by Xu et al., shows that NO has greater activity when the tumour is less<br />
differentiated and hence a higher grade.<br />
<br />
Using NO as a chemoattractant, the low and high levels of NO produced by cancer cells and macrophages, respectively can be perceived by the sensor which can target drugs directly to the tumour. An alternative method is to use oxygen as a chemorepellent so modified cells can specifically target hypoxic cells. Specificity can be increased through the use of targeting using specific peptide sequences which home onto receptors or glycoproteins which are specific to tumour cells such as the NGR peptide (Pasqualini, R. et al. 2000).<br />
<br />
<br><br><br><br><br><br><br><br><br><br />
<br />
=='''Diagnostics'''==<br />
<br />
[[File:E.chromi.jpg| 200px| left | thumbnail |This image is from the Cambridge E.chromi project, showing the coloured faecal matter as a form of diagnostic which has potential for identifying damage to the gut.]]NO is produced throughout the mammalian body in a range of cells, including endothelial cells (Bedioi.F et,al., 2002). NO has a range of functions within the body, from being a secondary messenger in signalling pathways to being a relaxing factor within the cardiovascular system (Pang, et,al.,2003). Abnormal levels of NO can be an indication of diseases and disorders such as hypertension, impotence and obesity (Bedioui.F et,al.,2002). A NO sensor can be augmented into two ways to aid diagnosis and treatment. For example, measuring the NO level of externally exhaled air can be an important and non-invasive diagnostic for airways inflammation. This is due to endothelial cells producing NO synthase (eNOS), both constitutive and inducible (iNOS). The iNOS is induced by pro inflammatory cytokines, as well as there being an increase in enzyme expression after exposure to oxidants. Therefore, measuring the NO level produced by eNOS has led to finding that there are higher levels of epithelial nitric oxide when asthma is untreated, and that there is a fall after anti-inflammatory treatment (Narang ,et al., 2002).<br />
<br />
Alternatively,Building upon the Cambridge 2009 iGEM E.Chromi project, the NO sensor can be incorporated to perceive abnormal levels of NO in the gut flora or damage to the gut, which can visually be seen in the form of colours in faecal matter (Cambridge iGEM team , 2009). For example NO levels increase during damage of the gut, potentially due to its role in functional repair ( Miller et al, 1993). Therefore,an NO sensor would produce particular colours that could correspond to different levels of NO expected for certain diseases and disorders.<br />
<br />
Another use is through the cells being stimulated to create a protein coat for resistance and persistence within the human body. These cells could be mass produced and contain vaccines which can be taken at birth. These cells can be used to target pathogens or other abnormalities that arise in the body.<br />
<br />
=='''Understanding Pathogens'''==<br />
<br />
[[File:Sensing.jpg| 200px| left | thumbnail |(Henares, et, al., 2012). The process used by bacteria in which NO course quorum sensing through lux U within V.harveyi.]]<br />
Pathogens have many different mechanisms and strategies of invasion and attack. For example some pathogens produce biofilms. To create biofilms, bacteria signal to other bacteria to assess the population size and density (Tsou, et al., 2009). When numbers are sufficient, they switch on gene expression which leads to aggregation and adhesion. This is an example of quorum sensing, as has been demonstrated through Nitric Oxide responsive quorum sensing circuits though lux U within Vibrio harveyi(Henares, et al., 2012). A greater understanding of NO can be applied to research into pathogenic diseases which could lead to advances in the creation of cures and vaccines.<br />
<br />
<br><br><br><br />
<br />
=='''Construction and Buildings Emissions'''==<br />
<br />
In 2008 the Modern Buildings Services released an article detailing recommendations from the Buildings Regulation Establishment Environmental Assessment Method (BREEAM) encouraging construction and buildings companies to monitor levels of nitric oxide emissions. They detail nitric oxide formation through combustion reactions involved in certain construction methods and even household applicances (e.g. boilers) and the consequences of excess atmospheric nitric oxide, including the production of smog and acid rain. Production of a way to accurately sense nitric oxide levels in the form of the hybrid promoter would be an extremely useful tool in detailing these levels; further to this combining the hybrid promoter and the comparator circuit could see bacteria that could not only sense nitric oxide being released by these appliances etc., but could also express an enzyme to safely break nitric oxide down and prevent the formation of harmful compounds.<br />
<br><br><br />
<br />
==References==<br />
<br />
Bedioui,F., Villeneuue, N.(2002)‘ Electrochemical Nitric Oxide Sensors for biological samples- Principle, selected examples and applications’, Electroanalysis, 15; 5-18. <br />
<br><br><br />
Cambridge iGEM team (2009) Available at: https://2009.igem.org/Team:Cambridge [Accessed on: 21/09.2012] <br />
<br><br><br />
Civerolo, K.L. and Dickerson, R.R. (1998) ‘Nitric oxide soil emissions from tilled and untilled corn®elds’, Agricultural and Forest Meteorology, 90; 307-311. <br />
<br><br><br />
Cole, C., Thomas, S., Filak, H., Henson, P., Lenz, L. (2012) ‘Nitric Oxide Increases Susceptibility of Toll-like Receptor-Activated Macrophages to Spreading Listeria monocytogenes’, Immunity, 36; 885.<br />
<br><br><br />
Davidson, E. (2012) Sources of Nitric Oxide and Nitrous Oxide following Wetting of Dry Soil, Soil Sci. Soc. Am. J. 56; 95–102<br />
<br><br><br />
Ferreira, L.F. and Behnke, B.J. (2010) ‘A toast to health and performance! Beetroot juice lowers blood pressure and the O2 cost of exercise’, Journal of Applied Physiology, 110; 585-586.<br />
<br><br><br />
Henares, B.M., Higgins, K.E., Boon, E.M. (2012)'Discovery of a Nitric Oxide Responsive Quorum Sensing Circuit in Vibrio harveyi',ACS chemical biology,17;1331-6.<br />
<br><br><br />
Lipschultz, F., Zafiriou, O.C. Wofsy, S.C., Elroy, M.B., Valois, F.W. and Watson, S.W. (1981) ‘Production of NO and N2O by soil nitrifying bacteria’, Macmillan Journals, 294; 641-643.<br />
<br><br><br />
Miller, M.J., Zhang, X.J., Sadowska-Krowicka, H., Chotinaruemol ,S., McIntyre ,J.A., Clark ,D.A. and Bustamante ,S.A. (1993) 'Nitric oxide release in response to gut injury',scandinavian journal of gastroenterology,28; 149-54.<br />
<br><br><br />
Modern Building Services, (2008), ''Understanding NOx emissions [online]'', Available at: [http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html http://www.modbs.co.uk/news/archivestory.php/aid/5380/Understanding_NOx_emissions.html], accessed on 26/09/2012<br />
<br><br><br />
Narang, I., Ersu,R., Wilson, N.M., Bush. A. (2002) 'Nitric oxide in chronic airway inflammation in children: diagnostic use and pathophysiological significance', Thorax ,57;586-589.<br />
<br><br><br />
Pang.J, Fan.C, Liv.X, Chem.T, Li.G (2003)'A nitric oxide biosensor or based on the multi-assembly of hemoglobin, montmorillonite/ polyvinyl alcohol at pyrolytic graphite electrode.', Bio sensors and bioelectronics, 19: 441-445. <br />
<br><br><br />
Pasqualini, R., Koivunen, E., Kain, R., Lahdenranta, J., Sakamoto, M., Stryhn, A., Ashmun, R.A., Shapiro, L.H., Arap, W. And Ruoslahti, E. (2000) ‘Aminopeptidase N is a receptor for tumour-homing peptides and a target for inhibiting angiogenesis’, The Journal of Cancer Research, 60; 722-727.<br />
<br><br><br />
Tsou,A.M., Cai,T., Liu,Z.,Zhu,J., and Kulkarni, R.V.,(2009)'Regulatory targets of quorum sensing in Vibrio cholerae: evidence for two distinct HapR-binding motifs',Nucleic Acids Res, 37; 2747–2756.<br />
<br><br><br />
Xu, W., Liu, L.Z., Loizidou, M., Ahmed, M. And Charles, I.G. (2002) ‘The role of nitric oxide in cancer’, Cell Research, 12; 311-320</div>RussellGritton