Team:Manchester/Notebooktest2

From 2013.igem.org

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      <a href="https://2013.igem.org/Team:Manchester/Hometest" id="link1">Home</a>
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      <a href="https://2013.igem.org/Team:Manchester/Hometest3" id="link1">Home</a>
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                   <li><a href="https://2013.igem.org/Team:Manchester/Modellingtest" id="link6">Modelling</a>
                   <li><a href="https://2013.igem.org/Team:Manchester/Modellingtest" id="link6">Modelling</a>
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    <li><a href="https://2013.igem.org/Team:Manchester/fattytest" id="link6">Fatty Acid Production</a></li>
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    <li><a href="https://2013.igem.org/Team:Manchester/parametertest" id="link6">Parameter Estimation</a></li>
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    <li><a href="https://2013.igem.org/Team:Manchester/parametertest" id="link6">Uncertainty Analysis</a></li>
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                            <li><a href="https://2013.igem.org/Team:Manchester/fabAmodeltest" id="link6">FabA Dynamics Model</a></li>
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                          <li><a href="https://2013.igem.org/Team:Manchester/popdynamictest" id="link6">Population Dynamics</a></li>
                             <li><a href="https://2013.igem.org/Team:Manchester/collabtest" id="link6">Modelling Collaboration</a></li>
                             <li><a href="https://2013.igem.org/Team:Manchester/collabtest" id="link6">Modelling Collaboration</a></li>
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<ul class="submenu2">
<ul class="submenu2">
  <li><a href="https://2013.igem.org/Team:Manchester/environmenttest" id="link4">Environmental Impact</a></li>
  <li><a href="https://2013.igem.org/Team:Manchester/environmenttest" id="link4">Environmental Impact</a></li>
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  <li><a href="https://2013.igem.org/Team:Manchester/economytest" id="link4">Economical Impact</a></li>
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  <li><a href="https://2013.igem.org/Team:Manchester/economytest" id="link4">Economic Impact</a></li>
  <li><a href="https://2013.igem.org/Team:Manchester/managementtest" id="link4">Impact Management</a></li>
  <li><a href="https://2013.igem.org/Team:Manchester/managementtest" id="link4">Impact Management</a></li>
  <li><a href="https://2013.igem.org/Team:Manchester/conclusiontest" id="link4">Conclusion</a></li>
  <li><a href="https://2013.igem.org/Team:Manchester/conclusiontest" id="link4">Conclusion</a></li>
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    <li><a href="https://2013.igem.org/Team:Manchester/businesstest" id="link4">Business Plan</a></li>
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                            <li><a href="https://2013.igem.org/Team:Manchester/businesstest" id="link4">Business Plan</a></li>
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    <li><a href="https://2013.igem.org/Team:Manchester/collabtest" id="link4">Modelling Collaboration</a></li>
                    <li><a href="https://2013.igem.org/Team:Manchester/knoledgetest" id="link4">Knowledge Deficit Assumption</a></li>
                    <li><a href="https://2013.igem.org/Team:Manchester/knoledgetest" id="link4">Knowledge Deficit Assumption</a></li>
                             <li><a href="https://2013.igem.org/Team:Manchester/conferencetest" id="link4">Conferences and Discussions</a></li>
                             <li><a href="https://2013.igem.org/Team:Manchester/conferencetest" id="link4">Conferences and Discussions</a></li>
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<li id="one"><a href="" onclick="blocking('text101'); return false;">Search for the Project</a>
<li id="one"><a href="" onclick="blocking('text101'); return false;">Search for the Project</a>
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Exams are over and we’ve officially started full-time work on the project! At the start of the week we had a big group meeting, complete with instructors and advisors, and were presented with our very own pins and stickers set. The pins quickly vanished, but we h
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<p>The team was put together in December ‘12 and work began right away to find a project for us to start working on. It quickly became apparent that you can’t will your imagination to start spouting brilliant ideas, but potential projects seemed to start appearing in everything we encountered in our day-to-day lives! Armed with this new source of ideas the team started meeting regularly throughout the week to have brainstorming sessions, where every idea, no matter how silly it seemed, was aired and judged by the rest of the group. Lots of pieces of paper later and we managed to whittle our ideas down into our three favourites.</p>
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<p>During the first few months of research we went to lots of conferences and discussions that were really useful (more info can be found on our Conferences and Discussions page). Firstly, going to meetings of these kind was great in terms of finding out what synthetic biology is currently used for, and the present outlooks for the area. Perhaps more importantly though, these meetings proved to be very good for networking with people who are either already involved with synthetic biology, or people who are interested in synthetic biology. This meant that we got some useful feedback on our budding ideas, which further aided their development.</p>
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<p>Around May, we had a major group meeting with Eriko and Rainer and most of our advisors. At this meeting we presented each of our three favourite ideas to the supervisors, and discussions followed surrounding the feasibility of each idea. Eventually the team reached a unanimous decision to power ahead with the synthetic alternative to palm oil components idea, and then the hard work started!</p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/c/c0/Rsz_searchproject.png"  /></center> </p>
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                 <li id="one"><a href="" onclick="blocking('text102'); return false;">Public Outreach Planning</a>
                 <li id="one"><a href="" onclick="blocking('text102'); return false;">Public Outreach Planning</a>
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Exams are over and we’ve officially started full-time work on the project! At the start of the week we had a big group meeting, complete with instructors and advisors, and were presented with our very own pins and stickers set. The pins quickly vanished, but we h
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<p>From the start of the project we had decided that we would include outreach activities aimed at young people in order to interest and educate them, but also to promote the field of synthetic biology to the next generation. Luckily, Elsa had some contacts within the Faculty of Life Sciences (FLS) at the university, and we quickly managed to sign up for both a two-day workshop event and the annual Community Open Day! (More info can be found on our Public Outreach pages)</p>
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<p>Having received word of a university-wide competition to encourage public engagement, Jess gave a presentation to the Head of Public Engagement within FLS and to the other competitors, ultimately winning us £150 to go towards our outreach events and an Outreach Mentor (Matthew Hickman)! We then met with Matthew and discussed our activity ideas, and he gave us lots of useful hints and tips of the dos and don’ts associated with hosting outreach events.</p>
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<p>After much deliberation we finally settled on what our main activity would be - a hands-on workshop where the children would build a DNA double helix out of sweets (representing the base pairs), strawberry pencils (representing the sugar-phosphate backbone) and cocktail sticks (representing the hydrogen bonding)! We also decided to include a mini discussion/debate amongst the children on the ethics of synthetic biology.</p>
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<p>During the weeks leading up to summer (and our planned events!), the outreach team designed a poster for the Community Open Day and made a thorough plan of how our workshops would be run. The aim of the poster was to attractively present our project, the palm oil industry and the iGEM competition to a wide range of people (the Open Day was free for anyone to attend), which we certainly did!</p>
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  <p>After realising just how expensive biology can be, we’ve started to make a big push on the sponsorship front. So far this has led to some free kits from QIAGEN, so we're off to a good start. </p>
  <p>After realising just how expensive biology can be, we’ve started to make a big push on the sponsorship front. So far this has led to some free kits from QIAGEN, so we're off to a good start. </p>
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  <p>Unfortunately this week we also had to say goodbye to Ali and Johanna, who need to spend time on other projects. Luckily we can say hello to Marco (who will be helping in the lab) and Denis (who’s now in charge of designing our wiki. It’s good to have a computer scientist on the team!). </p>
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  <p>Unfortunately this week we also had to say goodbye to Ali and Johanna, who need to spend time on other projects. Luckily we can say hello to Marco (who will be helping in the lab) and our new advisor Denis (who’s will be helping us out with implementing our wiki ideas). </p>
  <p>As you can see it’s been a fairly quiet week, but with our upcoming public outreach events and YSB 1.0, we’ve got a busy fortnight ahead of us!</p>
  <p>As you can see it’s been a fairly quiet week, but with our upcoming public outreach events and YSB 1.0, we’ve got a busy fortnight ahead of us!</p>
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            <li id="two"><a href="" onclick="blocking('text5'); return false;">Week 6</a>
            <li id="two"><a href="" onclick="blocking('text5'); return false;">Week 6</a>
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                               <p> This has been a very exciting week of collaborations for us! At the start of the week we had a Google hangout with Perdue to discuss their idea to standardise entry of parts into the registry. It emerged that there are many issues raised when it comes to standardising modelling. We agreed to help them with this. By the end of the week, this had somewhat grown...</p>
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                               <p> This has been a good week of collaborations for us! At the start of the week we had a Google hangout with Perdue to discuss their idea to standardise entry of parts into the registry. It emerged that there are many issues raised when it comes to standardising modelling. We agreed to help them with this. By the end of the week, this had somewhat grown...</p>
  <p>On Thursday we headed off to London for the very first Young Synthetic Biologists (YSB) conference. This was a big meeting of all the UK iGEM teams. On the first day we listened to presentations from each team in the morning. The afternoon featured workshops covering topics such as public engagement, business startups and bioart. </p>
  <p>On Thursday we headed off to London for the very first Young Synthetic Biologists (YSB) conference. This was a big meeting of all the UK iGEM teams. On the first day we listened to presentations from each team in the morning. The afternoon featured workshops covering topics such as public engagement, business startups and bioart. </p>
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  <p>The second day brought big news to the team. After our presentation, we had a lot of interest in our model. This led to a huge collaboration with 8 other UK teams:  Edinburgh, Westminster, Dundee, Newcastle, UCL, York and (maybe) Imperial. Our plan is two fold:</p>
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  <p>The second day brought exciting news to the team. After our presentation, we had a lot of interest in our model. This led to a proposed collaboration with several other UK teams. Our aim is to each make a video tutorial on how to use various different software commonly used to model in iGEM, and to then make these videos accessible to future teams. Hopefully having introductions like this will encourage and inspire more people to embrace modelling in their projects!</p>
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<p>1) Relating to the collaboration with Perdue, we are going to write some standardised pieces of code for use in the majority of the software. Also we would consider what essential pieces of information are needed for submission of a model. The idea being that one day we could have a standard registry for models. This is already being planned by iGEM, but we can use this project to provide feedback with what we'd want from it. </p>
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<p>2) An idea of a page that gives teams an introduction to the modelling on the iGEM website in the same way we have intros to the lab work. This would involve video tutorials on the main programmes used in iGEM. Other teams liked the idea and would consider doing one for the software they are using during the summer. Between the 8 teams we have experts in 10 different pieces of software!</p>
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<p>It appears big things could come from this project!</p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/a/a0/Ysb12.jpg"  /></center> </p>
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<p>At the end of the week we had another iGEM social where we tried the Icelandic delicacy of Hákarl (fermented shark). As you can imagine, it tasted bad but smelled even worse! The vegetable chilli that Jess made the team was much more appetising...</p>
<p>At the end of the week we had another iGEM social where we tried the Icelandic delicacy of Hákarl (fermented shark). As you can imagine, it tasted bad but smelled even worse! The vegetable chilli that Jess made the team was much more appetising...</p>
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<p><center>[Hain Daniels and Eccelso logos? (only if using Qiagen's from other week)]</center></p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/a/a8/Hakarl.jpg"  /></center> </p>
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<p>The project is getting increasingly stressful, which is seeing a rise in our team socialising time! Coincidence? I think not. This week the team visited Tim’s place for food on Saturday, then went to a few bars in town, our favourite of which was The Alchemist. If they serve drinks in conical flasks it still counts as research, right?</p>
<p>The project is getting increasingly stressful, which is seeing a rise in our team socialising time! Coincidence? I think not. This week the team visited Tim’s place for food on Saturday, then went to a few bars in town, our favourite of which was The Alchemist. If they serve drinks in conical flasks it still counts as research, right?</p>
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<p><center>[insert picture of Alchemist crew here]</center></p>
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The work was split into 4 sections: 1. Cloning and inserting ribosomal binding site (RBS) gene into pUC18 vector, 2. Cloning of FabA from WT <i>E. coli</i> BL21 (DE3), 3. FadD knockout, 4. LC-MS characterisation.</p>
The work was split into 4 sections: 1. Cloning and inserting ribosomal binding site (RBS) gene into pUC18 vector, 2. Cloning of FabA from WT <i>E. coli</i> BL21 (DE3), 3. FadD knockout, 4. LC-MS characterisation.</p>
<p>1. RBS BioBricks from the kit were ligated into plasmid, transformed into <i>E. coli</i> DH5-alpha (?), and miniprepped. (this is probably wrong. Rob or Tim clarify please)</p>
<p>1. RBS BioBricks from the kit were ligated into plasmid, transformed into <i>E. coli</i> DH5-alpha (?), and miniprepped. (this is probably wrong. Rob or Tim clarify please)</p>
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<p>2. We really want to BioBrick FabA because it is an important part of the fatty acid biosynthesis pathway of <i>E. coli</i> that is yet to be submitted to the Registry, and also we want to attempt to measure the kinetic properties of the enzyme to improve our model (include this?). This week we cloned the gene from <i>E. coli</i> BL21 (DE3) using PCR, extracted using gel electrophoresis and stored in the freezer. The team are now thinking on the best ways to characterise this gene.</p>
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<p>2. We really want to BioBrick FabA because it is an important part of the fatty acid biosynthesis pathway of <i>E. coli</i> that is yet to be submitted to the Registry, and also we would really like to attempt to measure the kinetic properties of the enzyme to improve our model. This week we cloned the gene from <i>E. coli</i> BL21 (DE3) using PCR and stored in the freezer. The team are now thinking on the best ways to characterise this gene.</p>
<p>3. Sadly, this week we had to say goodbye to our hopes of achieving knockout of the FadD gene. 10 long weeks and nothing to show for it (other than a significant improvement in our PCR and gel electrophoresis skills)! </p>
<p>3. Sadly, this week we had to say goodbye to our hopes of achieving knockout of the FadD gene. 10 long weeks and nothing to show for it (other than a significant improvement in our PCR and gel electrophoresis skills)! </p>
<p>4. We met with Nik who guided us through the quenching and metabolite extraction of prokaryotic cells. Initially we are running reference samples through Orbitrap LC-MS (WT <i>E. coli</i> BL21 (DE3) cells in different media, solid and liquid fractions of authentic palm oil), which we will then compare to the fatty acid profiles of our constructs. Nik also kept us entertained with stories about how the Orbitrap technology was developed in a cellar here in Manchester, and showed us a signed mass spectrometer by Alexander Makarov himself!</p>
<p>4. We met with Nik who guided us through the quenching and metabolite extraction of prokaryotic cells. Initially we are running reference samples through Orbitrap LC-MS (WT <i>E. coli</i> BL21 (DE3) cells in different media, solid and liquid fractions of authentic palm oil), which we will then compare to the fatty acid profiles of our constructs. Nik also kept us entertained with stories about how the Orbitrap technology was developed in a cellar here in Manchester, and showed us a signed mass spectrometer by Alexander Makarov himself!</p>
<p>This week’s social saw us at Jess’ house once more, this time for delicious Malaysian food and a film night after a particularly gruelling day in the lab. It goes without saying that everyone enjoyed letting their hair down for a bit.</p>
<p>This week’s social saw us at Jess’ house once more, this time for delicious Malaysian food and a film night after a particularly gruelling day in the lab. It goes without saying that everyone enjoyed letting their hair down for a bit.</p>
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<p><center>[insert picture of very busy whiteboard]</center></p>
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<p><center>[step 1. get picture of signed mass spec. -> step 2. put it here]</center></p>
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<p>Monday also saw Divita leave to go on holiday, so Tan (well, her sister!) cooked the team a great meal to say goodbye.</p>
<p>Monday also saw Divita leave to go on holiday, so Tan (well, her sister!) cooked the team a great meal to say goodbye.</p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/8/8c/Piclab5.jpg"  /></center> </p>
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        <li id="one"><a href="" onclick="blocking('text12'); return false;">Week 13</a>
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                               <p>This week has flown by! Partly because of how busy we were, but probably because it was a bank holiday on Monday so we’ve had a 4 day week (got to adhere to health and safety regulations!)</p>
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<p>Tuesday finally saw the arrival of our synthesised delta 9 and delta 12 genes, so the main objective of our project can finally begin!. Without any hesitation we hydrated them, transformed them into <i>E. coli </i>, and miniprepped them to get a lovely stock of DNA ready for us to use for a biobrick! We also inserted fabA in to a blunt-end vector and transformed it to get more DNA to work with. Next, they were all digested ready to ligate into the iGEM submission vector on Friday. Everything was going well right up to our awful gel extraction yields, meaning biobricks didn’t happen this week! It was very disappointing, but our weekend will be spent finding ways to improve our techniques.</p>
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<p> Once again this week we said goodbye to certain members of the team. Not one, not two, but THREE team members left us this week (obviously optimistic we’d have the project finished by now!). Now that Tim, Marco and Elsa are gone, we have 3 on the lab team and 2 on the modelling team.  With not long to go until the biobrick submission deadline, here’s hoping we work well under pressure! </p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/7/76/Piclab9.jpg"  /></center> </p>
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            <li id="two"><a href="" onclick="blocking('text13'); return false;">Week 14</a>
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                               <p>This week was filled with frustration, as our gel extractions refused to cooperate with us! Luckily however, Lorna soon became a gel extraction pro, and our DNA yield soon increased to workable levels. </p>
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<p>This week we also ordered our polo shirts, and we got the (correct!) sequencing results back from our fabA gene. </p>
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                               <p>It’s starting to feel like all of the effort we’ve been putting into the project will pay off! This week we managed to get our genes of interest (fabA, delta 9 and delta 12) into the iGEM submission plasmid! Sequencing showed that our genes are in fact ligated, and so we eagerly arranged a fedEX shipment to send our samples on their merry way to Boston. It’s not over though! We still needed to successfully ligate our genes into an expression plasmid. We chose pSB1C3 with a ribosomal binding site (RBS) and promoter (P) (Part BB1_ K608002). After a few seemingly failed attempts (our colonies were very small and took a while longer than expected to grow), we decided to test digest anyway because what did we have to lose (except all hope)? Excitingly however, the test digestions suggested that we’d put the genes into the expression plasmid! Time caught up with us and so we will be characterising next week. We think that the small colonies may be a result of the constitutive promoter used, and in an ideal world we’d check this theory. With one week to go though we will just have to leave it to any future iGEM teams out there!</p>
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<p>The pressure is very much on now, the whole team is feeling it. We’re desperately trying to juggle a chaotic lab schedule, writing up the wiki and making the presentation/poster for the jamboree.</p>
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<p> <center><img src="https://static.igem.org/mediawiki/2013/6/60/Piclab14.jpg"  /></center> </p>
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        <li id="four"><a href="" onclick="blocking('text15'); return false;">Week 16</a>
        <li id="four"><a href="" onclick="blocking('text15'); return false;">Week 16</a>
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                               <p>This is it! The final stretch to wikifreeze and the pressure is hotting up. At the start of the week we inoculated our RBS/P and Delta9/Delta12/fabA constructs in FAS media containing the fatty acids we needed to feed the bacteria with. We left our samples in the very capable hands of Dr Rattray who ran our samples on the Orbitrap LC-MS so we could see if our fatty acid profile had changed. The results finally came through the day before wikifreeze, and we were thrilled to find out  that our biobricks had worked! </p>
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<p>There’s no time to celebrate though. It’s the day of wiki freeze and we’re furiously working to get our wiki finished. Talk about cutting it fine!</p>
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                    <a href="https://2013.igem.org/Team:Manchester/Projecttest">PROJECT</a>
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Latest revision as of 17:22, 24 October 2013

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