View source for Team:Dundee/Lab Book/FluID

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            <h1><highlight class="highlight">FluID Detector</highlight></h1>
            <h3><highlight class="highlight">Lab Book</highlight></h3>
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          <a href="#section1" class="scroll"><span class="glyphicon glyphicon-briefcase" onclick="location.href='https://2015.igem.org/Team:Dundee/Lab_Book/FluID';" ></span></a> 
          <h3>FluID</h3>
          <p class="about-content">FluID is a cell free, all in one spray which will be used to detect and dinstiguish between body fluids found at crime scenes.</p>
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          <a href="#section2"><span class="glyphicon glyphicon-search" type="button" onclick="location.href='https://2015.igem.org/Team:Dundee/Lab_Book/Chromate_Detector';"></span></a>
          <h3>Chromate Biosensor</h3>
          <p class="about-content">Our chromate biosensor is a device that will be used to detect stainless steel residues left behind on bones in blunt or sharp force trauma cases.</p>
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          <a href="#section3"><span class="glyphicon glyphicon-eye-open" onclick="location.href='https://2015.igem.org/Team:Dundee/Lab_Book/Fingerprints';"></span></a> 
          <h3>Fingerprint Ageing</h3>
          <p class="about-content">This device will search for the presence of a specifc component found in fingerprint deposits whose presence/absence will indicate approximate fingerprint age.</p>
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        <div class="labtitle">Week Beginning 1/6/2015</div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"><i>Lactoferrin binding protein A (LbpA)</i> was successfully cloned into the vector pSB1C3.</p>
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            <span class="box-title">1/6: Amplification of <i>LbpA</i> for Cloning into pSB1C3</span>
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                  <p><b>Aim of Experiment:</b> Amplify <i>LbpA</i> from MC58 <i>N.meningitidis</i> template chromosome.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a> </p>
         <p><b>Results: </b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> Figure 1 </a></p> 
                  <p><b>Next Steps:</b> Since the <i>LbpA</i> gene was amplified successfully, it was ready to be digested for subsequent cloning into pSB1C3.</p>
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            <span class="box-title">2/6: Restriction Digests and Ligations with pSB1C3</span>
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                  <p><b>Aim of experiment:</b> To digest <i>LbpA</i> with BamHI and EcoRI and then ligate it into pSB1C3.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
         <p><b>Results:</b> N/A


                  <p><b>Next Steps:</b> Ligations of pSB1C3-<i>LbpA</i> were left overnight until the next day when they would be transformed into the JM110 <i>E.coli</i>.
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            <span class="box-title">3/6: Transformations of pSB1C3-<i>LpbA</i> into JM110 <i>E.coli</i> </span>
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                 <p><b>Aim of experiment:</b> To transform the pSB1C3-<i>LbpA</i> ligations into JM110 <i>E.coli</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a></p>
                  <p><b>Results:</b> N/A
                  <p><b>Next Steps:</b> Transformations will be checked tomorrow and if successful, overnight cultures will be set up.
                  
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            <span class="box-title">4/6: Overnight Cultures of JM110 <i>E.coli</i> containing pSB1C3-<i>LbpA</i></span>
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                 <p><b>Aim of experiment:</b> To set up overnight cultures of JM110 <i>E.coli</i> containing pSB1C3-<i>LbpA</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures
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                  <p><b>Results:</b> N/A
                  <p><b>Next Steps:</b> A miniprep will be performed tomorrow on the overnight cultures. 
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            <span class="box-title">5/6: Plasmid Purification of JM110 <i>E.coli</i> containing pSB1C3-<i>LbpA</i></span>
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                  <p><b>Aim of experiment:</b> To miniprep the JM110 <i>E.coli</i> overnight cultures containing pSB1C3-<i>LbpA</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit) </a>
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                  <p><b>Results:</b> N/A </p>
                  <p><b>Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing.                </div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Sequencing confirmed that <i>LbpA</i> had been successfully cloned into pSB1C3 therefore this week cloning was attempted to put <i>LbpA</i> into the high expression vector pQE80-L.</p>
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            <span class="box-title">8/6: Amplification of <i>LbpA</i> for Cloning into pQE80-L</span>
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                  <p><b>Aim of experiment:</b> To amplify <i>LbpA</i> for subsequent cloninng into the pQE80-L vector.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR </a></p>
                  <p><b>Results: </b>N/A</p>
<p><b> Next Steps:</b> The PCR product will be digested and ligated into pQE80-L tomorrrow.</p>
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            <span class="box-title">9/6: Restriction Digests and Ligations of <i>LbpA</i>into pQE80-L</span>
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                  <p><b>Aim of experiment:</b> To restrict and ligate the <i>LbpA</i> gene into the pQE80-L plasmid.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a> </p>
              <p><b> Results:</b>: N/A </p>
<p><b> Next Steps:</b> Transformations of pQE80-L-<i>LbpA</i> into JM110 <i>E.coli</i> will be carried out tomorrow.</p>
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            <span class="box-title">10/6: Transformations of pQE80-L-<i>LpbA</i> into JM110 <i>E.coli</i> </span>
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                  <p><b>Aim of experiment:</b> To transform  pQE80-L-<i>LpbA</i> into JM110 <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a></p>
              <p><b>Results:</b> N/A </p>
<p><b> Next Steps:</b> If the transformations are successful, colonies will be picked to set up overnight cultures. </p>
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        <div class="labtitle">Week Beginning 15/6/15</div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">This week, cloning of <i>LbpA</i> into pQE80-L was continued due to last week's failed attempts.</p>
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            <span class="box-title">15/6: PCR of <i>LbpA</i> for Cloning into pQE80-L</span>
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                  <p><b>Aim of experiment:</b> To re-amplify the <i>LbpA</i> gene for cloning into the pQE80-L vector.</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a></p>
                  <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure3-modal" class="journal-protocol">Figure 3</a></b> </p>
<p><b>Next Steps:</b> The gel shows that <i>LbpA</i> has been successfully amplified so it will be digested and ligated into the pQE80-L vector tomorrow.  </p>
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        <div class="labtitle">Week Beginning 22/6/15</div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"><i>LbpA</i> was successfully cloned into the high expression vector pQE80-L.</p>
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            <span class="box-title">23/6: Restriction Digest and Ligation of <i>LbpA</i> into pQE80-L</span>
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                  <p><b>Aim of experiment:</b> To digest the amplified <i>LbpA</i> gene with BamHI and KpnI and subsequently ligate it into the pQE80-L vector.</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The ligations will be transformed into M15[<i>pREP4</i>] <i>E.coli</i>. </p>

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            <span class="box-title">24/6: Transformations of pQE80-L-<i>LbpA</i> into M15[<i>pREP4</i>] <i>E.coli</i></span>
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                  <p><b>Aim of experiment:</b> To transform pQE80-L-<i>LbpA</i> into M15[<i>pREP4</i>] <i>E.coli</i>. </p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Overnight cultures will be set up tomorrow if the transformations are successful. </p>

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            <span class="box-title">25/6: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i> </span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Overnight cultures will be miniprepped and a pre-sequence restriction digest will be performed to check for the presence of the <i>LbpA</i> gene. </p>

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            <span class="box-title">26/6: Plasmid Purification of the M15[<i>pREP4</i>] <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>LbpA</i> </span>
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                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures from yesterday and perform a pre-sequencing restriction digest using BamHI and KpnI to check for presence of <i>LbpA</i> gene.</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification (QIAprep® Spin Miniprep Kit)</a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> A sample of the miniprep was sent for sequencing to confirm the presence of <i>LbpA</i> in the pQE80-L vector. </p>

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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Protein expression optimization experiments were performed on LbpA to assay protein production levels. However, cell growth seems to cease when <i>LbpA</i> expression is induced.</p>
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            <span class="box-title">1/7: M15[<i>pREP4</i>] <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>LbpA</i> </span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures in preparation for protein expression experiments tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a></p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The overnight cultures will be used to set up day cultures tomorrow which will be used to test expression of <i>LbpA</i>. </p>
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            <span class="box-title">2/7: Optimization of <i>LbpA</i> Expression</span>
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                  <p><b>Aim of experiment:</b> To set up day cultures and induce expression of <i>LbpA</i> using different concentrations of IPTG and check levels of protein production via SDS-PAGE.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization </a> </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The samples were stored in the -20 <sup>o</sup>C freezer overnight and will be ran on a gel tomorrow.</p>
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            <span class="box-title">3/7: Continuation of Optimization of <i>LbpA</i> Expression</span>
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                  <p><b>Aim of experiment:</b> To test the samples obtained yesterday on an SDS gel.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization </a> </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure4-modal" class="journal-protocol"> Figure 4 </a> </p>
<p><b>Next Steps:</b> Further optimization experiments will be set up next week using different concentrations of IPTG. </p>
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        <div class="labtitle">Week Beginning 6/7/15</div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Further protein expression optimization experiments were set up, this time using a range of IPTG concentrations to induce expression of <i>LbpA</i>.</p>
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            <span class="box-title">7/7: Further Optimization of <i>LbpA</i> Expression</span>
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                  <p><b>Aim of experiment:</b> To set up further tests to optimize expression of <i>LbpA</i> using different concentrations of IPTG.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization </a> Note: An SDS gel was not ran, instead the OD<sub>600</sub> readings were recorded for the uninduced and induced cultures at 20 minute intervals for 2 hours, 1 and half hours after the cultures were induced. </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure5-modal" class="journal-protocol"> Figure 5 </a> </p>
<p><b>Next Steps:</b> A plate reader experiment will be set up next week to monitor the growth rate of the the cells over a longer period of time after <i>LbpA</i> expression has been induced. </p>
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        <div class="labtitle">Week Beginning 13/7/15</div>
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          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Protein expression optimization experiments were continued this week. A growth curve assay was carried out to assess the effect of <i>LbpA</i> expression over a longer period of time.  </p>
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            <span class="box-title">14/7: Plate Reader Growth Curve Assay</span>
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                  <p><b>Aim of experiment:</b> To induce expression of <i>LbpA</i> using different concentration of IPTG and allow cells to grow for 16 hours in order to monitor cell growth.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#platereader-modal" class="journal-protocol"> Growth Curve Assay </a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure6-modal" class="journal-protocol"> Figure 6 </a> </p>
<p><b>Next Steps:</b> A plate reader experiment will be set up next week to monitor the growth rate of the the cells over a longer period of time after <i>LbpA</i> expression has been induced. </p>
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            <span class="box-title">17/7: Sample Preparation for Western Blot</span>
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                  <p><b>Aim of experiment:</b> To check if the cells in the induced cultures ~7 hours after being induced are producing LbpA. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot Sample Preparation </a> Note: The day cultures were grown for 7 hours after they were induced and then the samples were frozen, to be blotted on Monday. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The samples will be ran on an SDS gel on Monday and blotted. </p>
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<!-- WEEK 8 -->
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        <div class="labtitle">Week Beginning 20/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Samples from the growth curve assay were blotted to see if LbpA is being produced at the later stages of the growth curve. A further growth curve assay was also performed which monitored cell growth until a certain OD<sub>600</sub> was reached, at which point the cultures were induced.</p>
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            <span class="box-title">20/7: Western Blot LpbA Culture Samples</span>
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                  <p><b>Aim of experiment:</b> To western blot the LbpA culture samples from Friday.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot </a></p>
              <p><b>Results:</b> Nothing was visible on the blot. </p>
<p><b>Next Steps:</b> Another plate reader experiment will be set up on Thursday where the cultures will be monitored for three hours before induction with IPTG and then for a further 20 hours afterwards. </p>
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            <span class="box-title">23/7: Growth Curve Assay</span>
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                  <p><b>Aim of experiment:</b> To check if the cells begin to die after induction with IPTG after having already grown to an OD<sub>600</sub> of 0.5. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#platereader2-modal" class="journal-protocol"> Growth Curve Assay </a> </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure7-modal" class="journal-protocol"> Figure 7 </a> </p>
<p><b>Next Steps:</b> Samples of these cultures will be taken 6 hours after induction to check for the presence of LbpA. </p>
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<!-- WEEK 9 -->
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        <div class="labtitle">Week Beginning 27/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">The presence of LbpA was confirmed in the culture 6 hours after induction with IPTG so protein purification was started this week.</p>
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            <span class="box-title">27/7: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i> </span>
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                  <p><b>Aim of experiment:</b> To set up a 150 ml overnight culture for sub-culturing tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> Note: 150 ml of LB was used and 150 µl of each antibiotic. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> 150 ml of fresh LB will be inoculated tomorrow with 7.5 ml of the overnight cultures. </p>
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            <span class="box-title">28/7: <i>LbpA</i> Expression Assay</span>
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                  <p><b>Aim of experiment:</b> To set up day cultures using the overnight cultures from yesterday and test for expression of LbpA 6 hours after induction.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Test </a> Note: 150 ml of fresh LB was inoculated with 7.5 ml of the overnight cultures. Only an uninduced control and an 1 mM IPTG induced culture was set up. The samples were not blotted, they were only ran on an SDS gel. </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure8-modal" class="journal-protocol"> Figure 8 </a>   </p>
<p><b>Next Steps:</b> A 3 L culture will be set up on Thursday under the same conditions as the overnight cultures in order to try and purify LbpA.</p>
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            <span class="box-title">29/7: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i> for 3L Culture</span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures for the 3L day cultures that will be grown tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> Note: One 150 ml overnight culture was set up using 150 µl of each antibiotic. </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> The 3 L culture will be set up tomorrow allowed to grow to OD<sub>600</sub> of 0.6-1 and then induced with 1 mM IPTG and grown for 6 hours.</p>
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            <span class="box-title">30/7: 3 L Culture of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i> for Purification of LbpA</span>
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                  <p><b>Aim of experiment:</b> To set up 3 L day cultures for purifying LbpA.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The procedure was stopped at Step 6 and the pellets were frozen at -20 <sup>o</sup>C. </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> Protein purification will be continued on Monday.</p>
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<!-- WEEK 10 -->
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        <div class="labtitle">Week Beginning 3/8/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">The first round of LbpA purification failed at the affinity chromatography stage so another 6 L culture was grown this week to attempt purification again.</p>
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            <span class="box-title">3/8: Continuation of LbpA Purification</span>
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                  <p><b>Aim of experiment:</b> To continue with purification of LbpA.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The protocol was continued from Step 7</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>Size exclusion chromatography was not carried out since an SDS gel showed LbpA was not present after performing affinity chromatography. Another 6L culture will be set up this week.</p>
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            <span class="box-title">5/8: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#sacollapsiblew10-2"></span>
                <div id="sacollapsiblew10-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures for the 6 L culture that will be set up tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a>Note: One 150 ml overnight culture was set up using 150 µl of each antibiotic.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>The overnight cultures will be used to set up the 6 L day culture tomorrow. </p>
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            <span class="box-title">6/8: 6 L Day Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>LbpA</i></span>
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                <div id="sacollapsiblew10-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up 6 L day cultures for purifying LbpA and begin the purification process.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: The protocol was stopped at Step 6.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Purification of LbpA will be continued on Monday. </p>
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  </div>


<!-- WEEK 11 -->
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        <div class="labtitle">Week Beginning 10/8/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Purification of LbpA was continued but unfortunately was unsuccessful. If time permits, another round of purification may be reattempted. </p>
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            <span class="box-title">10/8: Continuation of LbpA Purification</span>
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                <div id="sacollapsiblew11-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To continue with purification of LbpA.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The protocol was continued from Step 7.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Purification of LbpA will be continued tomorrow. </p>
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            <span class="box-title">11/8: Continuation of LbpA Purification</span>
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                <div id="sacollapsiblew11-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To purify LbpA using affinity chromatography.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The protocol was stopped at Step 13. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>Since no peaks were observed, purification of LbpA was abadoned at this stage. Purification may be retried. </p>
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        </div>
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<!-- SEMEN-->
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        <span id="biospray-title">Semen</span>
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<!-- WEEK 1 -->
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        <div class="labtitle">Week Beginning 25/5/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i> PotD</i> into the biobrick vector, pSB1C3, was started this week.</p>
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            <span class="box-title">28/5: PCR of <i>PotD</i> from MG1655 <i>E.coli</i> gDNA</span>
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                  <p><b>Aim of Experiment:</b> To amplify <i>PotD</i> from MG1655 <i>E.coli</i> gDNA.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a></p>
         <p><b>Results: </b>N/A</p> 
                  <p><b>Next Steps:</b> The PCR product will be gel extracted, digested and ligated into the biobrick vector pSB1C3 tomorrow.</p>
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            <span class="box-title">29/5: Gel Extraction, Restriction Digest and Ligation of <i>SBP</i> into pSB1C3</span>
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                <div id="secollapsiblew1-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To gel extract the <i>PotD</i> PCR product and digest it with PstI and EcoRI. A ligation will then be set up with pSB1C3.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digest</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> Ligations will be transformed into JM110 <i>E.coli</i> on Monday.</p>
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<!-- WEEK 2 -->
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        <div class="labtitle">Week Beginning 1/6/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>PotD</i> into the biobrick vector was completed this week. </p>
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            <span class="box-title">1/6: Transformation of pSB1C3-<i>PotD</i> into JM110 <i>E.coli</i></span>
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                  <p><b>Aim of experiment:</b> To transform pSB1C3-<i>PotD</i> into JM110 <i>E.coli</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a></p>
                  <p><b>Results:</b>N/A</p>
<p><b> Next Steps:</b> If the transformations are successful, overnight cultures will be set up tomorrow.</p>
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            <span class="box-title">2/6: Overnight Cultures of pSB1C3-<i>PotD</i> in JM110 <i>E.coli</i></span>
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                <div id="secollapsiblew2-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of pSB1C3-<i>PotD</i> in JM110 <i>E.coli</i></p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a></p>
              <p><b> Results:</b>: N/A </p>
<p><b> Next Steps:</b> The overnight cultures will be miniprepped tomorrow. </p>
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            <span class="box-title">3/6: Plasmid Purification of pSB1C3-<i>PotD</i> from Overnight Cultures</span>
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                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures from yesterday.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit) </a></p>
              <p><b>Results:</b> N/A </p>
<p><b> Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing to confirm <i>PotD</i> has been inserted into pSB1C3 successfully.</p>
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<!-- WEEK 3 -->

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        <div class="labtitle">Week Beginning 15/6/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">This week, cloning of <i>PotD</i> into pQE80-L was started. The gene for <i>Spermine Binding Protein (SBP) </i> also arrived in a plasmid so it was transformed into MC1061 <i>E.coli</i> and plasmid purified.</p>
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            <span class="box-title">15/6: PCR of <i>PotD</i> for Cloning into pQE80-L</span>
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                  <p><b>Aim of experiment:</b> To amplify the <i>PotD</i> gene for cloning into the pQE80-L vector.</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a></p>
                  <p><b>Results:</b> <a data-toggle="modal" data-target="#lbpa-figure3-modal" class="journal-protocol">Figure 3</a></b> </p>
<p><b>Next Steps:</b> The gel shows that <i>PotD</i> has been successfully amplified so it will be digested and ligated into the pQE80-L vector tomorrow.  </p>
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        </div>
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            <span class="box-title">17/6: Restriction Digest and Ligation of <i>PotD</i> into pQE80-L. Transformation of IDT Plasmid containing <i>SBP</i> into MC1061 <i>E.coli</i></span>
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                <div id="secollapsiblew3-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest the <i>PotD</i> PCR product with KpnI and BamHI and then ligate it into pQE80-L. To transform MC1061 <i>E.coli</i> with the IDT plasmid containing <i>SBP</i>.</p>
                  <p><b>Protocols Used: </b></p><i>PotD</i>: <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a> <i>SBP</i>:<a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations</a> </p>
                  <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The pQE80-L-<i>PotD</i> ligations will be transformed into MC1061 <i>E.coli</i>. Overnight cultures of the transformations will be set up tomorrow.</p>
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            <span class="box-title">18/6: Transformations of <i>PotD</i> + pQE80-L into MC1061 <i>E.coli</i>. Overnight Cultures of MC1061 <i>E.coli</i> with SBP + IDT Plasmid</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#secollapsiblew3-4"></span>
                <div id="secollapsiblew3-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform the pQE80-L-<i>PotD</i> ligations into MC1061 <i>E.coli</i>. To set up overnight cultures of the transformations of MC1061 <i>E.coli</i> with <i>SBP</i> + IDT Plasmid that were set up yesterday.</p>
                  <p><b>Protocols Used: </b><i>PotD</i>:<a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a> <i>SBP</i>: <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures</a></p>
                  <p><b>Results:</b>N/A </p>
<p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up tomorrow.</p>
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            <span class="box-title">19/6: Repeat of Ligation of <i>PotD</i> into pQE80-L. Plasmid Purification of the IDT Plasmid Containing <i>SBP</i></span>
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                <div id="secollapsiblew3-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To repeat ligations of <i>PotD</i> into pQE80-L since transformations did not work. To miniprep the overnight cultures that were set up yesterday.</p>
                  <p><b>Protocols Used: </b><i>PotD</i>: <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a> <i>SBP</i>: <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Ligations will be left over the weekend and transformed on Monday. </p>
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<!-- WEEK 4 -->
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        <div class="labtitle">Week Beginning 22/6/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>PotD</i> into the high expression vector pQE80-L was completed. Cloning of <i>SBP</i> into pSB1C3 and pQE80-L was started this week.</p>
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         <div id="secollapsiblew4" class="collapse week-content"> 


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            <span class="box-title">22/6: Transformation of pQE80-L-<i>PotD</i> Ligations. PCR and Gel Extraction of <i>SBP</i> for Cloning into pQE80-L and Restriction Digests and Ligation of <i>SBP</i> for Cloning into pSB1C3</span>
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                  <p><b>Aim of Experiment:</b> To transform pQE80-L-<i>PotD</i> into  MC1061<i>E.coli</i>. To set up a PCR of <i>SBP</i> using the purified plasmid from Friday and to gel extract the PCR product for cloning into pQE80-L. To digest <i>SBP</i> out of the same plasmid using EcoRI and PstI and subsequently ligate it into pSB1C3.</p>
                  <p><b>Protocols Used:</b> <i>PotD</i>: <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a> <i>SBP</i>: <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
         <p><b>Results: </b><i>SBP</i> :<a data-toggle="modal" data-target="#sbp-figure1-modal" class="journal-protocol"> Figure 1 </a></p> 
                  <p><b>Next Steps:</b> Overnight cultures will be set up tomorrow for the <i>PotD</i> transformations. The <i>SBP</i> PCR product will be digested and ligated into pQE80-L tomorrow. The pSB1C3-<i>SBP</i> ligations will be transformed into MC1061 <i>E.coli</i>. </p>
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            <span class="box-title">23/6: Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>PotD</i>. Transformations of MC1061 <i>E.coli</i> with pSB1C3 + <i>SBP</i>. Restriction Digests and Ligations of <i>SBP</i> into pQE80-L and Transformations into MC1061 <i>E.coli</i> </span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures of MC1061 <i>E.coli</i> containing pSB1C3 + <i>PotD</i>. To transform MC1061 <i>E.coli</i> with pSB1C3 + <i>SBP</i>. To digest the <i>SBP</i> PCR product with BamHI and Kpn I, ligate it into pQE80-L and transform these ligations into MC1061 <i>E.coli</i>.</p>
                  <p><b>Protocols Used:</b> <i>PotD</i>: <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> <i>SBP</i>: <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a><a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The MC1061 <i>E.coli</i> overnight cultures containing pQE80-L + <i> PotD</i> will be miniiprepped tomorrow. Overnight cultures will be set up of the MC1061 <i>E.coli</i> transformations with pSB1C3 + <i>SBP</i> and pQE80-L + <i>SBP</i>.</p>
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            <span class="box-title">24/6: Plasmid Purification of the Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>PotD</i>. Overnight Cultures of the Transformations of MC1061 <i>E.coli</i> with pSB1C3 + <i>SBP</i> and pQE80-L + <i>SBP</i>.</span>
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                <div id="secollapsiblew4-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>PotD</i>. To set up overnight cultures of the transformations of MC1061 <i>E.coli</i> with pSB1C3-<i>SBP</i> and pQE80-L-<i>SBP</i></p>
                  <p><b>Protocols Used:</b> <i>PotD</i>: <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> <i>SBP</i>: <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with the highest concentration for pQE80-L + <i>PotD</i> was sent for sequencing. The <i>SBP</i> overnight cultures will be miniprepped tomorrow. </p>
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            <span class="box-title">25/6: Plasmid Purification of the Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i> and pQE80-L-<i>SBP</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#secollapsiblew4-4"></span>
                <div id="secollapsiblew4-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i> and pQE80-L + <i>SBP</i>.</p>
                  <p><b>Protocols Used:</b> <i>SBP</i>: <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit) </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with the highest concentration for both pSB1C3-<i>SBP</i> and pQE80-L-<i>SBP</i> were sent for sequencing. </p>
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<!-- WEEK 5 -->
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        <div class="labtitle">Week Beginning 29/6/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Sequencing showed <i> SBP</i> failed to insert successfully into pSB1C3 so cloning was repeated from the ligation stage this week. However, SBP and PotD were successfully inserted into the high expression vector pQE80-L, so overexpression assays were started this week for both.</p>
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            <span class="box-title">29/6: Ligations of <i>SBP</i> into pSB1C3 and Transformations into MC1061 <i>E.coli</i> </span>
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                <div id="secollapsiblew5-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To ligate <i>SBP</i> into pSB1C3 and transform the ligations into MC1061 <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <i>SBP</i>: <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a></p>
         <p><b>Results: </b><i>SBP</i> :<a data-toggle="modal" data-target="#sbp-figure1-modal" class="journal-protocol"> Figure 1 </a></p> 
                  <p><b>Next Steps:</b> Overnight cultures will be set up tomorrow if transformations are successful. </p>
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            <span class="box-title">30/6: Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i>, pQE80-L-<i>SBP</i> and pQE80-L-<i>PotD</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#secollapsiblew5-2"></span>
                <div id="secollapsiblew5-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i>. To set up overnight cultures  of pQE80-L-<i>SBP</i> and pQE80-L + <i>PotD</i> in MC1061 <i>E.coli</i> for overexpression assays to be performed tomorrow. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The MC1061 <i>E.coli</i> overnight cultures containing pSB1C3-<i> SBP</i> will be miniiprepped tomorrow. The MC1061 <i>E.coli</i> overnight cultures containing pQE80-L-<i> SBP</i> and pQE80-L-<i>PotD</i> will be subcultured tomorrow in order to perform overexpression assays.</p>
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            <span class="box-title">1/7: Plasmid Purification of the Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i>. <i>PotD</i> and <i>SBP</i> Expression Optimization</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#secollapsiblew5-3"></span>
                <div id="secollapsiblew5-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>SBP</i>. To start protein expression optimization for PotD and SBP.</p>
                  <p><b>Protocols Used:</b> <i>SBP</i>: <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> <i>SBP</i> and <i>PotD</i>: <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization</a> Note: Protocol was stopped at Step 8. Samples will be ran on an SDS gel tomorrow.</p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with the highest concentration for pSB1C3 + <i>SBP</i> was sent for sequencing. The PotD and SBP samples will be ran on an SDS gel tomorrow. </p>
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            <span class="box-title">2/7: Continuation of <i>PotD</i> and <i>SBP</i> Expression Optimization</span>
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                <div id="secollapsiblew5-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To run the samples of PotD and SBP on an SDS gel and begin Western Blotting.</p>
                  <p><b>Protocols Used:</b> <i>PotD</i> and <i>SBP</i>: <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization</a> Note: Protocol was continued from Step 9.</p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure2-modal" class="journal-protocol"> Figure 2 </a></p>
       <p><b>Next Steps:</b> The membrane for the Western blot has been left at the blocking stage so will be continued tomorrow.</p>
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            <span class="box-title">3/7: Western Blot of <i>PotD</i> and <i>SBP</i> Day Culture Samples</span>
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                <div id="secollapsiblew5-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To continue the Western Blot of PotD and SBP.</p>
                  <p><b>Protocols Used:</b> <i>PotD</i> and <i>SBP</i>: <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot</a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure3-modal" class="journal-protocol"> Figure 3 </a></p>
       <p><b>Next Steps:</b> PotD has been successfully characterized so purification of this protein will be started next week. Further expression optimization experiments for SBP will be set up next week using different concentrations of IPTG, since its characterization was unsuccessful.</p>
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<!-- WEEK 6 -->
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        <div class="labtitle">Week Beginning 6/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Further protein expression optimization experiments were set up, this time using a range of IPTG concentrations to induce expression of <i>SBP</i>.</p>
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            <span class="box-title">6/7: Further Optimization of <i>SBP</i> Expression</span>
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                  <p><b>Aim of experiment:</b> To set up further tests to optimize expression of <i>SBP</i> using different concentrations of IPTG.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization </a> Note: Cultures were induced with 0.5 mM, 1 mM and 2 mM IPTG. The Western Blot was left to block overnight. </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure4-modal" class="journal-protocol"> Figure 4 </a> </p>
<p><b>Next Steps:</b> The Western Blot will be completed tomorrow.</p>
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            <span class="box-title">7/7: Western Blot of <i>SBP</i> </span>
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                <div id="secollapsiblew6-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To complete the Western Blot that was started yesterday. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot </a> Note: Protocol was continued from Step 12.</p>
              <p><b>Results:</b> Nothing was visible on the blot. </p>
<p><b>Next Steps:</b> A plate reader experiment will be set up next week to monitor the growth rate of the the cells over a longer period of time after <i>SBP</i> expression has been induced.  </p>
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      </div> 
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<!-- WEEK 7 -->
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        <div class="labtitle">Week Beginning 13/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Protein expression optimization experiments were continued this week. A growth curve assay was carried out to assess the effect of <i>SBP</i> expression over a longer period of time. Purification of <i>PotD</i> was also started this week. </p>
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            <span class="box-title">14/7: Plate Reader Growth Curve Assay</span>
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                  <p><b>Aim of experiment:</b> To induce expression of <i>SBP</i> using different concentration of IPTG and allow cells to grow for 16 hours in order to monitor cell growth.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#platereader-modal" class="journal-protocol"> Growth Curve Assay </a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure5-modal" class="journal-protocol"> Figure 5 </a> </p>
<p><b>Next Steps:</b> A plate reader experiment will be set up next week to monitor the growth rate of the the cells over a longer period of time after <i>SBP</i> expression has been induced. </p>
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            <span class="box-title">15/7: Overnight Cultures for 3 L Culture for PotD Purification</span>
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                <div id="secollapsiblew7-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures that will be used tomorrow to set up 3 L cultures for PotD purification. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a></p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The 3 L day culture will be set up tomorrow.</p>
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            <span class="box-title">16/7: Sample Preparation for Western Blot of SBP and 3 L Day Culture for PotD Purification</span>
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                  <p><b>Aim of experiment:</b> To check if the cells in the induced cultures ~7 hours after being induced are producing SBP. To set up 3 L Day Culture for <i> PotD</i> Purification </p>
                  <p><b>Protocols Used: </b> <i>SBP</i>: <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot Sample Preparation </a> Note: The day cultures were grown for 7 hours after they were induced and then the samples were frozen, to be blotted on Monday. <i>PotD</i>:  <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a>Note: Cultures were left to grow overnight after being induced. </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The samples will be ran on an SDS gel on Monday and blotted. The <i>PotD</i> cultures will be harvested tomorrow. </p>
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            <span class="box-title">17/7: Harvesting <i>PotD</i> Cultures</span>
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                  <p><b>Aim of experiment:</b> To harvest the day cultures grown yesterday for <i>PotD</i> purification. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: After the cultures were spun down the pellets were frozen. </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> Purification of <i>PotD</i> will continue on Monday.  </p>
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<!-- WEEK 8 -->
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        <div class="labtitle">Week Beginning 20/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Samples from the growth curve assay were blotted to see if SBP is being produced at the later stages of the growth curve. A further growth curve assay was also performed which monitored cell growth until a certain OD<sub>600</sub> was reached, at which point the cultures were induced. Purification of PotD was also continued this week.</p>
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            <span class="box-title">20/7: Western Blot <i>SBP</i> Culture Samples</span>
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                  <p><b>Aim of experiment:</b> To western blot the SBP culture samples from Friday. To continue purfying PotD.</p>
                  <p><b>Protocols Used: </b> <i>SBP</i>: <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot </a>                           <i>PotD</i>: <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: Affinity chromatography was completed but no peaks were observable. </p>
              <p><b>Results:</b> Nothing was visible on the blot. </p>
<p><b>Next Steps:</b> Another plate reader experiment will be set up on Thursday where the cultures will be monitored for three hours before induction with IPTG and then for a further 20 hours afterwards. Another 150 ml overnight culture was set up for <i>PotD</i> so 3 L cultures could be set up tomorrow. </p>
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            <span class="box-title">21/7: 3 L Day Cultures for PotD Purification</span>
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                  <p><b>Aim of experiment:</b> To set up 3 L Day Culture for PotD Purification. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: Cultures were left to grow overnight after being induced. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Purification will be continued tomorrow. </p>
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            <span class="box-title">22/7: Continuation of PotD Purification </span>
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                  <p><b>Aim of experiment:</b> To continue purification of PotD. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: The protocol was continued from Step 4 and stopped at Step 13.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Purification will be continued tomorrow. </p>
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            <span class="box-title">23/7: Continuation of PotD Purification and Growth Curve Assay for <i>SBP</i> </span>
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                  <p><b>Aim of experiment:</b> To continue purification of PotD and to check if the cells begin to die after inducing expression of <i>SBP</i> with IPTG after having already grown to an OD<sub>600</sub> of 0.5. </p>
                  <p><b>Protocols Used: </b> <i>PotD</i>: <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: The protocol was continued from Step 13. <i>SBP</i>: <a data-toggle="modal" data-target="#platereader2-modal" class="journal-protocol"> Growth Curve Assay </a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure6-modal" class="journal-protocol"> Figure 6 </a> </p>
<p><b>Next Steps:</b> The samples from the size exclusion chromatography performed on PotD will be prepared tomorrow for blotting next week. For <i> SBP</i> further overexpression tests will be set up next week.</p>
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            <span class="box-title">24/7: SDS-PAGE and Western Blot of PotD  </span>
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                  <p><b>Aim of experiment:</b> To run SDS-PAGE of the elutions obtained on Friday after SEC.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> SDS-PAGE</a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure7-modal" class="journal-protocol"> Figure 7 </a> </p>
<p><b>Next Steps:</b> The same samples will be Western Blotted on Monday to confirm that the bands oberved are PotD.</p>
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<!-- WEEK 9 -->
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        <div class="labtitle">Week Beginning 27/7/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">PotD was successfully purified and characterized this week. SBP was still unobservable in induced 150ml cultures so 3 L cultures containing pQE80-L-<i>SBP</i> were set up. </p>
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            <span class="box-title">27/7: Overnight Cultures for <i>SBP</i> Expression Assays and SDS-PAGE and Western Blot of PotD</span>
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                  <p><b>Aim of experiment:</b> To set up a 150 ml overnight culture containing pQE80-L-<i>SBP</i> for sub-culturing tomorrow and to run an SDS-PAGE and Western Blot the PotD samples from Friday.</p>
                  <p><b>Protocols Used: </b><i>SBP</i>: <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> Note: 150 ml of LB was used and 150 µl of each antibiotic. <i>PotD</i>: <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> SDS-PAGE and Western Blotting</a> Note: The membrane was left blocking overnight.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> 150 ml of fresh LB will be inoculated tomorrow with 7.5 ml of the overnight cultures containing pQE80-L-<i>SBP</i>. The PotD Western Blot will be completed tomorrow. </p>
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            <span class="box-title">28/7: <i>SBP</i> Expression Assay and Completion of PotD Westen Blot</span>
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                  <p><b>Aim of experiment:</b> To set up day cultures using the overnight cultures from yesterday and test for expression of LbpA 6 hours after induction.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Test </a> Note: 150 ml of fresh LB was innoculated with 7.5ml of the overnight cultures. Only an uninduced control and an 1 mM IPTG induced culture was set up. The samples were not blotted, they were only ran on an SDS gel. </p>
              <p><b>Results:</b> <i>SBP</i>: Nothing was observable on the SDS gel. <i>PotD</i>: <a data-toggle="modal" data-target="#sbp-figure8-modal" class="journal-protocol"> Figure 8 </a> </p>
<p><b>Next Steps:</b> Since PotD has now been successful purified, the next step is to attach a fluorescent nanobead to the protein. Hopefully, this will be attempted in the coming weeks. Even though SBP was not visible on the SDS gel, 3 L cultures will be set up this week to try and harvest the protein since the culture size may be too small. </p>
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            <span class="box-title">29/7: Overnight Cultures for 3L Culture</span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures containing pQE80-L-<i>SBP</i> for the 3L day cultures that will be grown tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> Note: One 150ml overnight culture was set up using 150µl of each antibiotic. </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> The 3L culture will be set up tomorrow allowed to grow to OD<sub>600</sub> of 0.6-1 and then <i>SBP</i> expression induced with 1mM IPTG and grown for 6 hours.</p>
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            <span class="box-title">30/7: 3 L Culture for Purification of SBP</span>
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                  <p><b>Aim of experiment:</b> To set up 3 L day cultures for purifying SBP.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The procedure was stopped at Step 6 and the pellets were frozen at -20<sup>o</sup>C. </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> Protein purification will be continued on Monday.</p>
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  <!-- WEEK 10 -->
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        <div class="labtitle">Week Beginning 3/8/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">It seems that SBP has been successfully purified and characterized. However, there was still a lot of contamination in the samples after size eclusion chromatography was performed. .</p>
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            <span class="box-title">3/8: Continuation of SBP Purification</span>
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                  <p><b>Aim of experiment:</b> To continue with purification of SBP.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The protocol was continued from Step 7</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Size exclusion chromatography will be carried out tomorrow. </p>
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            <span class="box-title">4/8: Continuation of SBP Purification</span>
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                  <p><b>Aim of experiment:</b> To continue purification of SBP.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a>Note: The protocol was continued from Step 14. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The fractions corresponding to the peak observed during SEC, will be ran on an SDS gel and Western Blotted tomorrow. </p>
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            <span class="box-title">6/8: Western Blot of SBP</span>
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                <div id="secollapsiblew10-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To perform a Western Blot on the samples obtained from Size Exclusion Chromatography yesterday..</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blot </a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#sbp-figure9-modal" class="journal-protocol"> Figure 9 </a> </p>
<p><b>Next Steps:</b> There was still some contamination in the gel filtrated samples so this will have to be removed before a nanobead can be attached to the protein. </b> </p>
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  <!-- WEEK 11 -->
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        <div class="labtitle">Week Beginning 10/8/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Interactions between PotD and spermidine were analysed via tryptophan fluorescence this week.</p>
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            <span class="box-title">10/8: Tryptophan Fluorescence</span>
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                  <p><b>Aim of experiment:</b> To analyse the interaction of PotD and spermidine using tryptophan fluorescence. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#tf-modal" class="journal-protocol"> Tryptophan Fluorescence</a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b></p>
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        </div>
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<!-- BLOOD-->
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        <span id="biospray-title">Blood</span>
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<!-- WEEK 1 -->
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        <div class="labtitle">Week Beginning 10/6/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBA</i> and <i>hHBB</i> into the biobrick vector, pSB1C3, was performed this week.</p>
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            <span class="box-title">10/6: Restriction Digests of <i>hHBA</i> and <i>hHBB</i> from IDT gBlocks and Ligation into pSB1C3</span>
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                  <p><b>Aim of Experiment:</b> To digest <i>hHBA</i> and <i>hHBB</i> from IDT gBlocks using EcoRI and PstI for ligation into pSB1C3.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
         <p><b>Results: </b>N/A</p> 
                  <p><b>Next Steps:</b> The pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> ligations will be transformed into JM110 <i>E.coli</i>. </p>
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            <span class="box-title">11/6: Transformations of pSB1C3-hHBA and pSB1C3-hHBB Ligations into JM110 <i>E.coli</i></span>
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                  <p><b>Aim of experiment:</b> To transform the pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> ligations into JM1110 <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up.</p>
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<!-- WEEK 2 -->
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        <div class="labtitle">Week Beginning 15/6/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">The sequence for pSB1C3-<i>hHBB</i> was confirmed this week so cloning of <i>hHBB</i> into the two hybrid system vector pUT18 was started. Sequencing of pSB1C3-<i>hHBA</i> was incorrect so another miniprep was sent for sequencing.</p>
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            <span class="box-title">15/6: Overnight Cultures of JM110 <i>E.coli</i> containing pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i></span>
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                  <p><b>Aim of experiment:</b> To set up overnight cultures of JM110 <i>E.coli</i> containing pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> from the transformations performed on Tuesday.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> Overnight cultures will be miniprepped tomorrow and a test restriction digest will be performed to check for the presence of the inserts.</p>
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            <span class="box-title">16/6: Plasmid Purification of the JM110 <i>E.coli</i> Overnight Cultures containing pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> </span>
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                  <p><b>Aim of experiment:</b> To miniprep the JM110 <i>E.coli</i> overnight cultures containing pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> and perform a pre-sequence digest check.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification (QIAprep® Spin Miniprep Kit) </a> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests</a> </p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure1-modal" class="journal-protocol"> Figure 1 </a>
       <p><b>Next Steps:</b> The gel indicates that <i>hHBA</i> and <i>hHBB</i> have inserted successfully into pSB1C3 so samples of the minipreps will be sent for sequencing to confirm this.</p>
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            <span class="box-title">17/6: Amplification of <i>hHBA</i> and <i>hHBB</i> for Cloning into the Two Hybrid System Vectors </span>
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                  <p><b>Aim of experiment:</b> To amplify <i>hHBA</i> and <i>hHBB</i> for Cloning into the Two Hybrid System Vectors pT25 and pUT18, respectively. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure2-modal" class="journal-protocol"> Figure 2 </a>
       <p><b>Next Steps:</b> Sequencing of pSB1C3-<i>hHBA</i> and pSB1C3-<i>hHBB</i> showed that pSB1c3-<i>hHBA</i> had a 1 bp deletion so one of the other minipreps was sent for sequencing. pSB1C3-<i>hHBB</i>, however, was successful so the band for <i>hHBB</i> will be gel extracted and digested tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">18/6: Gel Extraction and Restriction Digests <i>hHBB</i> for Cloning into pUT18 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew2-4"></span>
                <div id="blcollapsiblew2-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To gel extract and digest <i>hHBB</i> with BamHI and EcoRI.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> Ligations of <i>hHBB</i> will be set up tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">19/6: Ligations of <i>hHBB</i> into pUT18</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew2-5"></span>
                <div id="blcollapsiblew2-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up ligations of pUT18-<i>hHBB</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> Ligations will be left over the weekend and transformed into BTH101 <i>E.coli</i>. </p>
                </div>
        </div>
        </div>
      </div> 
         </div>
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   </div>
  </div>


<!-- WEEK 3 -->

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    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 22/6/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Cloning of <i>hHBA</i> and <i>hHBB</i> into pT25 and pUT18, respectively, was continued this week.</p>
         </div>

         <div id="blcollapsiblew3" class="collapse week-content"> 


     <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">22/6: Transformations of pUT18-<i>hHBB</i> into BTH101 <i>E.coli</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew3-1"></span>
                <div id="blcollapsiblew3-1" class="collapse box-content">
                  <p><b> Aim of experiment:</b> To transform pUT18-<i>hHBB</i> into BTH101 <i>E.coli</i>.</p>
                  <p><b> Protocols Used: </b><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a></p>
                  <p><b> Results:</b>N/A </p>
<p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up tomorrow. Sequencing showed the sequence for pSB1C3-<i>hHBA</i> was incorrect again, a third miniprep will be sent for sequencing.</p>
</div>
        </div>
        </div>
      </div>  

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">23/6: Overnight Cultures of BTH101 <i>E.coli</i> containing pUT18-<i>hHBB</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew3-2"></span>
                <div id="blcollapsiblew3-2" class="collapse box-content">
                  <p><b> Aim of experiment:</b> To set up overnight cultures of BTH101 <i>E.coli</i> containing pUT18-<i>hHBB</i> from the transformations carried out yesterday.</p>
                  <p><b> Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures</a></p>
                  <p><b> Results:</b> N/A </p>
<p><b>Next Steps:</b> The overnight cultures will be miniprepped and sent for sequencing tomorrow. </p>
</div>
        </div>
        </div>
      </div>       

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">24/6: Plasmid Purification of the BTH101 <i>E.coli</i> Overnight Cultures containing pUT18-<i>hHBB</i></i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew3-3"></span>
                <div id="blcollapsiblew3-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the BTH101 <i>E.coli</i> Overnight Cultures containing pUT18-<i>hHBB</i>.</p>
                  <p><b>Protocols Used: </b></p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a></p>
                  <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing to confirm presence of insert. Sequencing for pSB1C3-<i>hHBA</i> came back correct so another PCR of <i>hHBA</i> will be set up tomorrow for cloning into pT25. </p>
</div>
        </div>
        </div>
      </div>  

      <div class="row">
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        <div class="box">
            <span class="box-title">25/6: Amplification of <i>hHBA</i> for Cloning into pT25</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew3-4"></span>
                <div id="blcollapsiblew3-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To amplify <i>hHBA</i> for cloning into pT25</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a></p>
                  <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure3-modal" class="journal-protocol"> Figure 3 </a> </p>
<p><b>Next Steps:</b> The PCR product digested and ligated into pT25 tomorrow.</p>
</div>
        </div>
        </div>
      </div>
      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">26/6: Restriction Digests and Ligations of <i>hHBA</i> for Cloning into pT25</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew3-5"></span>
                <div id="blcollapsiblew3-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest the <i>hHBA</i> PCR product with BamHI and KpnI and set up ligations with pT25.</p>
                  <p><b>Protocols Used: </b><a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests</a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a></p>
                  <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Ligations will be left over the weekend and transformed into BTH101 <i>E.coli</i> on Monday. </p>
</div>
        </div>
        </div>
      </div>

         </div>
     </div>
   </div>
  </div>



<!-- WEEK 4 -->
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    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 29/6/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew4"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBA</i> into pT25 was completed this week. Sequencing confirmed that <i>hHBB</i> has been successfully inserted into pUT18. Cloning of both <i>hHBA</i> and <i>hHBB</i>, individually, into the high expression vector pQE80-L was started this week.</p>
         </div>

         <div id="blcollapsiblew4" class="collapse week-content"> 


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">29/6: Transformation of pT25-<i>hHBA</i> Ligations into BTH101 <i>E.coli</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew4-1"></span>
                <div id="blcollapsiblew4-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To transform pT25-<i>hHBA</i> into  BTH101 <i>E.coli</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a></p>
         <p><b>Results: </b>N/A</p> 
                  <p><b>Next Steps:</b> Overnight cultures will be set up tomorrow for the pT25-<i>hHBA</i> transformations if successful. Sequencing confirmed <i>hHBB</i> has been successfully inserted into pUT18. </p>
                </div>
        </div>
        </div>
      </div>


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">30/6: Overnight Cultures of MC1061 <i>E.coli</i> containing pUT18-<i>hHBA</i> and Amplification of <i>hHBA</i> and <i>hHBB</i> for Cloning into pQE80-L</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew4-2"></span>
                <div id="blcollapsiblew4-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MC1061 <i>E.coli</i> containing pT25-<i>hHBA</i>. To amplify <i>hHBA</i>.and <i>hHBB</i> for cloning into pQE80-L.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a>  <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a><a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure4-modal" class="journal-protocol"> Figure 4 </a> </p>
       <p><b>Next Steps:</b> The BTH101 <i>E.coli</i> overnight cultures containing pT25-<i>hHBB</i> will be miniiprepped tomorrow. The PCR products will be digested and ligate dinto pQE80-L tomorrow.</p>
                </div>
        </div>
        </div>
      </div>


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">1/7: Plasmid Purification of the Overnight Cultures of BTH101 <i>E.coli</i> containing pT25-<i>hHBA</i> and Restriction Digests and Ligations of <i>hHBA</i> and <i>hHBB</i> into pQE80-L</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew4-3"></span>
                <div id="blcollapsiblew4-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the overnight cultures of BTH101 <i>E.coli</i> containing pT25-<i>hHBA</i>. To digest <i>hHBA</i> and <i>hHBB</i> with BamHI and KpnI and set up ligations with pQE80-L.</p>
                  <p><b>Protocols Used:</b><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a> </p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with the highest concentration for pT25-<i>hHBA</i> will be sent for sequencing. The pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i> will be transformed into M15[<i>pREP4</i>] <i>E.coli</i> tomrorrow.</p> 
                </div>
        </div>
        </div>
      </div>
      

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">2/7: Transformations of pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i> Ligations into M15[<i>pREP4</i>] <i>E.coli</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew4-4"></span>
                <div id="blcollapsiblew4-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform the pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i> ligations into M15[<i>pREP4</i>] <i>E.coli</i>.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up on Monday. </p>
                </div>
        </div>
        </div>
      </div>      
         </div>
     </div>
   </div>
  </div>


<!-- WEEK 5 -->
<div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 6/7/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBA</i> and <i>hHBB</i> into pQE80-L was completed this week. Cloning of <i>Haptoglobin (hHBN) </i> was also started this week.</p>
         </div>

         <div id="blcollapsiblew5" class="collapse week-content"> 


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">6/7: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5-1"></span>
                <div id="blcollapsiblew5-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To set up overnight cultures of BTH101 <i>E.coli</i> containing pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> </p>
         <p><b>Results: </b> N/A </p> 
      <p><b>Next Steps:</b>The overnight cultures will be miniprepped tomorrow. Sequencing for pT25-<i>hHBA</i> was incorrect so cloning will be repeated from the restriction digest stage.</p>
                </div>
        </div>
        </div>
      </div>


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">7/7: Plasmid Purification of M15[<i>pREP4</i>] <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5-2"></span>
                <div id="blcollapsiblew5-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up miniprep the BTH101 <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">  Plasmid Purification (QIAprep® Spin Miniprep Kit) </a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure5-modal" class="journal-protocol"> Figure 5</a> </p>
       <p><b>Next Steps:</b> The pre-sequence digest indicates that both <i>hHBA</i> and <i>hHBB</i> have been inserted into pQE80-L successfully. The minipreps with the highest concentration will be sent for sequencing. The sequence for <i>Haptoglobin (hHBN)</i> arrived in an IDT plasmid, work will be be started on this tomorrow.</p>
                </div>
        </div>
        </div>
      </div>


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">8/7: Transformation of IDT Plasmid-<i>hHBN</i> into MC1061 <i>E.coli</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5-3"></span>
                <div id="blcollapsiblew5-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform the IDT Plasmid-<i>hHBN</i> into MC1061 <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations</a> </p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> Overnight cultures will be set up tomorrow. </div>
        </div>
        </div>
      </div>
      

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">9/7: Overnight Cultures of MC1061 <i>E.coli</i> containing IDT Plasmid-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5-4"></span>
                <div id="blcollapsiblew5-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MC1061 <i>E.coli</i> containing IDT Plasmid-<i>hHBN</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures</a> </p>
         <p><b>Results:</b> N/A</p>
       <p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow and <i>hHBN</i> will be amplified for cloning into pSB1C3. </p>
                </div>
        </div>
        </div>
      </div>  
     

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">10/7: Plasmid Purification of IDT Plasmid-<i>hHBN</i> and Digestion of <i>hHBN</i> for Cloning into pSB1C3</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew5-5"></span>
                <div id="blcollapsiblew5-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the MC1061 <i>E.coli</i> containing IDT Plasmid-<i>hHBN</i> and to digest <i>hHBN</i> out of the plasmid using EcoRI and PstI.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests</a> </p>
         <p><b>Results:</b> N/A</p>
       <p><b>Next Steps:</b> <i>hHBN</i> will be ligated into pSB1C3 on Monday. Sequencing also confirmed <i>hHBA</i> and <i>hHBB</i> have been successfully inserted into pQE80-L.</p>
                </div>
        </div>
        </div>
      </div>     
         </div>
     </div>
   </div>
  </div>


<!-- WEEK 6 -->
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    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 13/7/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">hHBB was successfully characterized this week so protein purification was started. Cloning of <i>hHBN</i> into pSB1C3 was also completed.</p>
         </div>

         <div id="blcollapsiblew6" class="collapse week-content">
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">13/7: Ligations of <i>hHBN</i> into pSB1C3 and Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBA</i> and pQE80-L-<i>hHBB</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6-1"></span>
                <div id="blcollapsiblew6-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To ligate <i>hHBN</i> into pSB1C3. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures</a> </p>
              <p><b>Results:</b>N/A</p>
<p><b>Next Steps:</b> The pSB1C3-<i>hHBN</i> ligations will be transformed into MC1061 <i>E.coli</i> tomorrow. The overnight cultures will be subcultured tomorrow and induced with IPTG to test protein expression levels. </p>
</div>
        </div>
        </div>
      </div> 

<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">14/7: Transformation of pSB1C3-<i>hHBN</i> into MC1061 <i>E.coli</i> and Optmiization of <i>hHBA</i> and <i>hHBB</i> Expression </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6-2"></span>
                <div id="blcollapsiblew6-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform pSB1C3-<i>hHBN</i> into MC1061 <i>E.coli</i> and set up expression optimization tests for <i>hHBA</i> and <i>hHBB</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Protein Expression Optimization </a> </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure6-modal" class="journal-protocol"> Figure 6 </a></p>
<p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up tomorrow. Western Blotting will be continued tomorrow for <i>hHBA</i> and <i>hHBB</i>. </p>
</div>
        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">15/7: Overnight Cultures of MC1061 <i>E.coli</i> containing pSB1C3-<i>hHBN</i> and Western Blotting of <i>hHBA</i> and <i>hHBB</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6-3"></span>
                <div id="blcollapsiblew6-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight culture of MC1061 <i>E.coli</i> containing pSB1C3-<i>hHBN</i> and finish Western Blotting of <i>hHBA</i> and <i>hHBB</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blotting</a> </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure7-modal" class="journal-protocol"> Figure 7 </a> </p>
<p><b>Next Steps:</b> The blot shows that characterization of hHBB has been successful. Unfortunately, nothing was visible on the blot for hHBA. 3 L cultures for purification of hHBB will be set up this week. Characterization of hHBA may be reattempted. The overnight cultures will be miniprepped tomorrow.</p>
</div>
        </div>
        </div>
      </div> 

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        <div class="box">
            <span class="box-title">16/7: Plasmid Purification of pSB1C3-<i>hHBN</i> and Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBB</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6-4"></span>
                <div id="blcollapsiblew6-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the MC1061 <i>E.coli</i> overnight cultures containing pSB1C3-<i>hHBN</i> and set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBB</i> for 3L cultures. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing. The overnight cultures will be used to set up 3L day cultures tomorrow for purrification of hHBB. </p>
</div>
        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">17/7: 3 L Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBB</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew6-5"></span>
                <div id="blcollapsiblew6-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up 3 L day cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBB</i> for purification of hHBB. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: The protocol was stopped at Step 6. </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> Purification will be continued on Monday.</p>
</div>
        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>

<!-- WEEK 7 -->
<div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 20/7/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">hHBB was successfully purified this week and cloning of <i>hHBN</i> into pQE80-L was completed. </p>
         </div>

         <div id="blcollapsiblew7" class="collapse week-content">
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       <div class="border-day">
        <div class="box">
            <span class="box-title">20/7: Amplification of <i>hHBN</i> for Cloning into pQE80-L and Purification of hHBB</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7-1"></span>
                <div id="blcollapsiblew7-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To amplify <i>hHBN</i> for cloning into pQE80-L and continuation of hHBB purification.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR </a> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: Protocol was continued from Step 6. </p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure8-modal" class="journal-protocol"> Figure 8</a> </p>
<p><b>Next Steps:</b> The fractions from the affinity chromatography will be consolidated and concentrated tomorrow. Size exclusion chromatography will also be performed. The <i>hHBN</i> PCR product will be digested and ligated into pQE80-L tomorrow. </p>
</div>
        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">21/7: Size Exclusion Chromatography of hHBB and Restriction Digests and Ligations of <i>hHBN</i> into pQE80-L</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7-2"></span>
                <div id="blcollapsiblew7-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To consolidate, concentrate and perform size exclusion chromatography on the hHBB samples. To digest the <i>hHBN</i> PCR product with BamHI and KpnI and subsequently ligate it into pQE80-L. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> SDS </a> Note: An SDS gel was ran of the SEC fractions which corresponded to the observed peak. Western blotting was stopped at the blocking stage. <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a></p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure9-modal" class="journal-protocol"> Figure 9 </a> </p>
<p><b>Next Steps:</b> Western blotting will be completed tomorrow and will confirm if the bands observed on the SDS gel are indeed hHBB. The pQE80-L<i>hHBN</i> ligations will be transformed into M15[<i>pREP4</i>] <i>E.coli</i> tomorrrow. </p>
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        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">22/7: Western Blot of hHBB and Transformations of M15[<i>pREP4</i>] <i>E.coli</i> with pQE80-L<i>hHBN</i> Ligations</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7-3"></span>
                <div id="blcollapsiblew7-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To complete the Western blot of hHBB and transform pQE80-L<i>hHBN</i> into M15[<i>pREP4</i>] <i>E.coli</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blotting </a> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a>  </p>
              <p><b>Results:</b><a data-toggle="modal" data-target="#bl-figure10-modal" class="journal-protocol"> Figure 10 </a></p>
<p><b>Next Steps:</b> The blot shows hHBB has been successfully purified. If the transformations are successful, overnight cultures will be set up tomorrow. </p>
</div>
        </div>
        </div>
      </div>

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       <div class="border-day">
        <div class="box">
            <span class="box-title">23/7: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L<i>hHBN</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7-4"></span>
                <div id="blcollapsiblew7-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L<i>hHBN</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow.</p>
</div>
        </div>
        </div>
      </div>

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       <div class="border-day">
        <div class="box">
            <span class="box-title">24/7: Plasmid Purification of Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L<i>hHBN</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew7-5"></span>
                <div id="blcollapsiblew7-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep M15[<i>pREP4</i>] <i>E.coli</i> overnight cultures containing pQE80-L<i>hHBN</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing. </p>
</div>
        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>

<!-- WEEK 8 -->
<div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 27/7/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"><i>hHBN</i> was successfully cloned into pQE80-L and cloning started for insertion into pT25.</p>
         </div>

         <div id="blcollapsiblew8" class="collapse week-content">
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       <div class="border-day">
        <div class="box">
            <span class="box-title">27/7: Restriction Digests and Ligations of <i>hHBN</i> into pQE80-L</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8-1"></span>
                <div id="blcollapsiblew8-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest <i>hHBN</i> again with BamHI and KpnI and set up ligations with pQE80-L.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Resriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations </a> </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> Ligations will be transformed into M15[<i>pREP4</i>] <i>E.coli</i> tomorrow. </p>
</div>
        </div>
        </div>
      </div>

<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">28/7: Transformation of M15[<i>pREP4</i>] <i>E.coli</i> with pQE80-L-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8-2"></span>
                <div id="blcollapsiblew8-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform M15[<i>pREP4</i>] <i>E.coli</i> with pQE80-L-<i>hHBN</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a></p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up tomorrow. </p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">29/7: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i>  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8-3"></span>
                <div id="blcollapsiblew8-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i> </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow. </p>
</div>
        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">30/7: Plasmid Purification of M15[<i>pREP4</i>] <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8-4"></span>
                <div id="blcollapsiblew8-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the M15[<i>pREP4</i>] <i>E.coli</i> Overnight Cultures containing pQE80-L-<i>hHBN</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>The miniprep with the highest concentration will be sent for sequencing. </p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">31/7: Amplification of <i>hHBN</i> for Cloning into pT25</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew8-5"></span>
                <div id="blcollapsiblew8-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To amplfy <i>hHBN</i> for cloning into the two hybrid vector pT25.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a></p>
              <p><b>Results:</b><a data-toggle="modal" data-target="#bl-figure15-modal" class="journal-protocol"> Figure 11 </a> </p>
<p><b>Next Steps:</b>The PCR product will be digested and ligated into pT25 on Monday.</p>
</div>
        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>

<!-- WEEK 9 -->
<div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 3/8/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew9"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBN</i> into pT25 was completed this week but subsequent sequencing showed that it had not inserted successfully. </p>
         </div>

         <div id="blcollapsiblew9" class="collapse week-content">
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">3/8: Restriction Digests and Ligations of <i>hHBN</i> for Cloning into pT25</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew9-1"></span>
                <div id="blcollapsiblew9-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest the <i>hHBN</i> PCR product with BamHI and KpnI and set up ligations with the pT25 vector.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol"> Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligations</a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>The pT25-<i>hHBN</i> ligations will be transformed into MG1655 <i>E.coli</i> tomorrow. </p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">4/8: Transformations of  pT25-<i>hHBN</i> into MG1655 <i>E.coli</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew9-2"></span>
                <div id="blcollapsiblew9-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up transformations of pT25-<i>hHBN</i> into MG1655 <i>E.coli</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformations </a> </p>
              <p><b>Results:</b> N/A</p>
<p><b>Next Steps:</b> If transformations are successful, overnight cultures will be set up tomorrow. </p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">5/8: Overnight Cultures of MG1655 <i>E.coli</i> containing pT25-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew9-3"></span>
                <div id="blcollapsiblew9-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MG1655 <i>E.coli</i> containing pT25-<i>hHBN</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a> </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow. </p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">6/8: Plasmid Purification of MG1655 <i>E.coli</i> Overnight Cultures containing pT25-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew9-4"></span>
                <div id="blcollapsiblew9-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the  MG1655 <i>E.coli</i> overnight cultures containing pT25-<i>hHBN</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification (QIAprep® Spin Miniprep Kit)</a> </p>
              <p><b>Results:</b> N/A   </p>
<p><b>Next Steps:</b> The miniprep with the highest concentration will be sent for sequencing. </p>
</div>
        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>


  <!-- WEEK 10 -->
<div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 10/8/15</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew10"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Overexpression assays of <i>hHBN</i> were set up this week and purification of the protein was begun. </p>
         </div>

         <div id="blcollapsiblew10" class="collapse week-content">
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">10/8: Overnight Cultures for <i>hHBN</i> Overexpression Assays</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew10-1"></span>
                <div id="blcollapsiblew10-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i>. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures </a></p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b>The overnight cultures will be used to set up day cultures that will be induced with different concentrations of IPTG to assay <i>hHBN</i> expression levels.</p>
</div>
        </div>
        </div>
      </div>
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">11/8: Overexpression Assay of pQE80-L-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew10-2"></span>
                <div id="blcollapsiblew10-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To assay <i>hHBN</i> expression levels when induced with a range of IPTG concentrations. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Overexpression Assay </a>Note: Day cultures were grown to an OD<sub>600</sub> of 0.6 and then induced with 0.25 mM, 0.5 mM, 0.75 mM and 1 mM IPTG and allowed to grow for 6 hours. </p>
              <p><b>Results:</b> SDS gel </p>
<p><b>Next Steps:</b> The gel indicates that <i>hHBN</i> is not being overexpressed in any of the cultures. Further optimization experiments will be set up to try and overexpress <i>hHBN</i>. </p>
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        </div>
        </div>
      </div>
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            <span class="box-title">12/8: Overnight Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew10-3"></span>
                <div id="blcollapsiblew10-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i> which will be used to set up a 3 L day culture tomorrow.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol"> Overnight Cultures</a> Note: A 150 ml overnight culture was set up. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The overnight cultures will be used to set up 3 L day cultures tomorrow.  </p>
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        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">13/8: 3 L Day Cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew10-4"></span>
                <div id="blcollapsiblew10-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up 3 L day cultures of M15[<i>pREP4</i>] <i>E.coli</i> containing pQE80-L-<i>hHBN</i> using the overnight culture set up yesterday.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> Note: The cultures were grown to OD<sub>600</sub>=0.9, then induced with 1 mM IPTG and allowed to grow for a further 8 hours. A 1 ml sample was taken before the cultures were spun down. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> Protein purification will be continued next week. </p>
</div>
        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>

  <!-- WEEK 11 -->
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     <div class="box">
        <div class="labtitle">Week Beginning 17/8/15</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Purification of haptoglobin was completed this week and confirmed via Western Blotting. </p>
         </div>

         <div id="blcollapsiblew11" class="collapse week-content">
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">17/8: hHBN Purification</span>
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                <div id="blcollapsiblew11-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To continue purification of hHBN by performing affinity chromatography. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification </a> Note: The protocol was stopped at Step 13.</p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The samples corresponding to the peak observed in the chromatogram will be combined and purified using SEC tomorrow. </p>
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        </div>
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      </div>
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        <div class="box">
            <span class="box-title">18/8: hHBN Purification </span>
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                <div id="blcollapsiblew11-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To continue purification of hHBN by performing size exclusion chromatography. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#pp-modal" class="journal-protocol"> Protein Purification</a> </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps</b>: The gel shows bands corresponding to 45 kDa, the expected size of hHBN. This will be checked via Western Blotting to ensure it is the correct protein. </p>
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        </div>
        </div>
      </div>
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            <span class="box-title">19/8: Western Blotting of Purified hHBN</span>
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                <div id="blcollapsiblew11-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To perform Western Blotting in order to verify the protein that was purified is indeed hHBN. </p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blotting </a> Note: The membrane was left blocking in milk overnight. </p>
              <p><b>Results:</b> N/A </p>
<p><b>Next Steps:</b> The blot will be completed tomorrow. </p>
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        </div>
        </div>
      </div>

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        <div class="box">
            <span class="box-title">20/8: Western Blotting of Purified hHBN</span>
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                <div id="blcollapsiblew11-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To complete Western Blotting of hHBN.</p>
                  <p><b>Protocols Used: </b> <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> Western Blotting</a> The protocol was continued from Step 12.</p>
              <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure14-modal" class="journal-protocol"> Figure 14</a> </p>
<p><b>Next Steps:</b> The blot indicates that hHBN has been successfully purified.</p>
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        </div>
        </div>
      </div>
         </div>
     </div>
   </div>
  </div>


<!-- WEEK 12 -->
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        <div class="labtitle">Week Beginning 24/8/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBA</i> into pT25 was carried out this week. </p>
         </div>
         <div id="blcollapsiblew12" class="collapse week-content">


      <div class="row">
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        <div class="box">
            <span class="box-title">24/8: Amplification of <i>hHBA</i> for Cloning into pT25</span>
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                <div id="blcollapsiblew12-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To amplify <i>hHBA</i> for subsequent cloning into the two hybrid vector pT25.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure16-modal" class="journal-protocol"> Figure 15</a>
       <p><b>Next Steps:</b> The gel extracted PCR product will be digested and ligated into pT25 tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
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        <div class="box">
            <span class="box-title">25/8: Restriction Digests of <i>hHBA</i> and Ligations into pT25 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew12-2"></span>
                <div id="blcollapsiblew12-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest the <i>hHBA</i> with BamHI and KpnI and set up ligations with pT25. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligations</a> </p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b>The ligations will be left overnight and transformed into MG1065∆cya <i>E.coli</i> tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
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        <div class="box">
            <span class="box-title">26/8: Transformations of pT25-<i>hHBA</i> into MG1065∆cya <i>E.coli</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew12-3"></span>
                <div id="blcollapsiblew12-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform the pT25-<i>hHBA</i> ligations into MG1065∆cya <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> If transformations are successful, colonies will be pick for setting up overnight cultures tomorrow.  </p>
                </div>
        </div>
        </div>
      </div>

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       <div class="border-day">
        <div class="box">
            <span class="box-title">27/8: Overnight Cultures of MG1065∆cya <i>E.coli</i> containing pT25-<i>hHBA</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew12-4"></span>
                <div id="blcollapsiblew12-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MG1065∆cya <i>E.coli</i> containing pT25-<i>hHBA</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow.</p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">28/8: Plasmid Purification of MG1065∆cya <i>E.coli</i> containing pT25-<i>hHBA</i> Overnight Cultures</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew12-5"></span>
                <div id="blcollapsiblew12-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the  MG1065∆cya <i>E.coli</i> overnight cultures containing pT25-<i>hHBA</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification (QIAprep® Spin Miniprep Kit)</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with highest concentration was sent for sequencing to confirm if the insert has been cloned in successfully. </p>
                </div>
        </div>
        </div>
      </div> 
         </div>
     </div>
   </div>
  </div>


<!-- WEEK 13 -->
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     <div class="box">
        <div class="labtitle">Week Beginning 7/9/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Cloning of <i>hHBN</i> into pUT18 was carried out this week. </p>
         </div>
         <div id="blcollapsiblew13" class="collapse week-content">


      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">7/9: Amplification of <i>hHBN</i> for Cloning into pUT18</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew13-1"></span>
                <div id="blcollapsiblew13-1" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To amplify <i>hHBN</i> for subsequent cloning into the two hybrid vector pUT18.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#bl-figure17-modal" class="journal-protocol"> Figure 16</a> </p>
       <p><b>Next Steps:</b> The gel extracted PCR product will be digested and ligated into pT25 tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">8/9: Restriction Digests of <i>hHBN</i> and Ligations into pUT18 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew13-2"></span>
                <div id="blcollapsiblew13-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To digest the <i>hHBN</i> with BamHI and KpnI and set up ligations with pUT18. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests </a> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligations</a> </p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b>The ligations will be left overnight and transformed into MG1065∆cya <i>E.coli</i> tomorrow. </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">9/9: Transformations of pUT18-<i>hHBN</i> into MG1065∆cya <i>E.coli</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew13-3"></span>
                <div id="blcollapsiblew13-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To transform the pUT18-<i>hHBN</i> ligations into MG1065∆cya <i>E.coli</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations </a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> If transformations are successful, colonies will be picked for setting up overnight cultures tomorrow.  </p>
                </div>
        </div>
        </div>
      </div>

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">10/9: Overnight Cultures of MG1065∆cya <i>E.coli</i> containing pUT18-<i>hHBN</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew13-4"></span>
                <div id="blcollapsiblew13-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To set up overnight cultures of MG1065∆cya <i>E.coli</i> containing pUT18-<i>hHBN</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The overnight cultures will be miniprepped tomorrow.</p>
                </div>
        </div>
        </div>
      </div>

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       <div class="border-day">
        <div class="box">
            <span class="box-title">11/9: Plasmid Purification of MG1065∆cya <i>E.coli</i> containing pUT18-<i>hHBN</i> Overnight Cultures</span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#blcollapsiblew13-5"></span>
                <div id="blcollapsiblew13-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To miniprep the  MG1065∆cya <i>E.coli</i> overnight cultures containing pUT18-<i>hHBN</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification (QIAprep® Spin Miniprep Kit)</a></p>
         <p><b>Results:</b> N/A
       <p><b>Next Steps:</b> The miniprep with highest concentration was sent for sequencing to confirm if the insert has been cloned in successfully. </p>
                </div>
        </div>
        </div>
      </div> 
         </div>
     </div>
   </div>
  </div>


         </div>
       </div>
   </div>
</div>

  <!-- Nasal Mucus -->
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        <span id="biospray-title">Nasal Mucus </span>
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           <div id="nmcollapsible" class="collapse biospray-content">




<!-- WEEK 1 -->
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     <div class="box">
        <div class="labtitle">Week Beginning 15/6/2015</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew1"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Produced a purified plasmid of pIDT-<i>OBP2A</i> in order to use it as template in PCR. </p>
          </div>
        <div id="nmcollapsiblew1" class="collapse week-content">

      <div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">17/6: Transformation of pIDT-<i>OBP2A</i> in MC1610 <i>E.coli</i> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew1-1"></span>
                <div id="nmcollapsiblew1-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To transform pIDT-<i>OBP2A</i> into MC1061 <i> e.coli</i> strain. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> The result of this transformation can be seen tomorrow. If successful then the next step will be to produce an overnight culture in preparation of subsequent plasmid purification. </p>
                </div>
        </div>
        </div>
      </div>


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        <div class="box">
            <span class="box-title">18/6: Overnight culture of pIDT-<i>OBP2A</i> plasmid in MC1061  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew1-2"></span>
                <div id="nmcollapsiblew1-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To produce an overnight culture of pIDT-<i>OBP2A</i> from positive colonies of MC1061 from the transformation done on the 17/6.  
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Culture </a></p>
         <p><b>Results:</b> N/A
                  <p><b>Next Steps:</b> If the overnight culture successfully grows then the next stage of experimentation will be to produce a plasmid purification of pIDT-<i>OBP2A</i> from that overnight culture.                </div>
        </div>
        </div>
      </div>


      <div class="row">
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        <div class="box">
            <span class="box-title">19/6: Plasmid Purification of pIDT-<i>OBP2A</i> from the overnight culture    </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew1-3"></span>
                <div id="nmcollapsiblew1-3" class="collapse box-content">
                 <p><b>Aim of experiment:</b> The overnight culture produced on the 18/6 will now undergo a plasmid purification in order to obtain  the pIDT-<I>OBP2A</I> plasmid.</p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid Purification </a></p>
                  <p><b>Results:</b> The plasmid purification yielded a concentration of 305.53 ng/ul.
                  <p><b>Next Steps:</b> Assuming the plasmid purification has been successful, then the next stage of experimentation will be to run a PCR using this purified pIDT-<i>OBP2A</i> plasmid as the template for subsequent amplification.              </div>
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        </div>
      </div>


    </div>
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  </div>


                  
     

<!-- WEEK 2 -->
  <div class="row">
    <div class="border-week">
        <div class="labtitle">Week Beginning 22/6/2015</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Removal of the signaling peptide from <i>OBP2A</i> and insertion of  <i>OBP2A</i> into the biobrick vector pSB1C3. </p>
         </div>
         <div id="nmcollapsiblew2" class="collapse week-content">

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        <div class="box">
            <span class="box-title">22/6: PCR of pIDT-<I>OBP2A</I> using primers designed for the removal of the signal peptide  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2-1"></span>
                <div id="nmcollapsiblew2-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a PCR of the purified pIDT-<I>OBP2A</I> plasmid using a set of primers that have been designed to remove the signaling peptide at the start of the <I>OBP2A</I> gene, whilst leaving the rest of the gene intact.  </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> If the PCR is successful, then the next step will be to gel extract <i>OBP2A</i> from a  the  gel corresponding to the size of the <I>OBP2A</I> gene fragment.  </p>
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        </div>
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            <span class="box-title">23/6: Gel extraction of <I>OBP2A</I> from the PCR performed on the 22/6  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2-2"></span>
                <div id="nmcollapsiblew2-2" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To perform a gel extraction of <i>OBP2A</i> from a PCR reaction that was performed on the 22/6. Once gel extraction has been performed a subsequent gel will be done to determine if the gel extraction had been successful.   
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol">Gel Extraction </a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#nm-figure1-modal" class="journal-protocol"> Figure 1 </a><a data-toggle="modal" data-target="#nm-figure2-modal" class="journal-protocol"> Figure 2</a>

 <p><b>Next Steps:</b> To perform a restriction of <I>OBP2A</I> in preparation for ligation.             
 </div>
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        <div class="box">
            <span class="box-title">24/6:  Restriction, ligation and transformation of <I>OBP2A</I> into pSB1C3 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2-3"></span>
                <div id="nmcollapsiblew2-3" class="collapse box-content">
                  <p><b>Aim of experiment:</b>  To perform a restriction, ligation and subsequently a transformation of  of the gel extracted <I>OBP2A</I> into the JM110 <I>E.coli</I> strain.   
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digest  </a>:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation </a>:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation  </a></p>
         <p><b>Results:</b> N/A


                  <p><b>Next Steps:</b> If the transformation is successful and positive colonies form then these colonies will then be grown overnight in preparation for plasmid purification.                </div>
        </div>
        </div>
      </div>


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        <div class="box">
            <span class="box-title">25/6: Re-ligation and transformation of <I>OBP2A</I> into pSB1C3 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2-4"></span>
                <div id="nmcollapsiblew2-4" class="collapse box-content">
                  <p><b>Aim of experiment:</b> The transformation performed on the 24/6 failed and as a result <I>OBP2A</I> will now be re-ligated into the biobrick vector pSB1C3 and subsequently transformed into the JM110 <I>E.coli</I> strain.   
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#nm-figure3-modal" class="journal-protocol"> Figure 3 </a>


                  <p><b>Next Steps:</b>If the transformation is successful and positive colonies form then these colonies will then be grown overnight in preparation for plasmid purification.                   </div>
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        </div>
      </div>


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            <span class="box-title">26/6: Overnight cultures of pSB1C3-<I>OBP2A</I> </span>
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                <div id="nmcollapsiblew2-5" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To produce overnight cultures of pSB1C3-<i>OBP2A</i> from a positive colony from the transformation done on the 25/6.  
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a></p>
         <p><b>Results:</b> N/A</p> 

                  <p><b>Next Steps:</b>  Produce plasmid purifications from the overnight cultures of pSB1C3-<i>OBP2A</i> in preparation for presequence digest.                </div>
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        </div>
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        <div class="box">
            <span class="box-title">27/6: Plasmid Purification and Presequence Digest of pSB1C3-<I>OBP2A</I>  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew2-6"></span>
                <div id="nmcollapsiblew2-6" class="collapse box-content">
                  <p><b>Aim of experiment:</b> To perform a plasmid purification of the overnight cultures of pSB1C3-<i>OBP2A</i> from the 26/6. Then to subsequently perform a presequence digest of those plasmid purification.   
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Restriction Digests</a></p>
         <p><b>Results: </b> <a data-toggle="modal" data-target="#nm-figure4-modal" class="journal-protocol"> Figure 4 </a>

               <p><b>Next Steps:</b> It was concluded from the results of these experiments that sample 5 of pSB1C3-<i>OBP2A</i> will be sent for sequencing.
                    </div>
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<!-- WEEK 3 -->
  <div class="row">
    <div class="box">
        <div class="labtitle">Week Beginning 29/6/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Positive Sequencing of pSB1C3-<I>OBP2A</I> and subsequent plasmid purification of pSB1C3-<i>OBP2A</i>. </p>
         </div>
         <div id="nmcollapsiblew3" class="collapse week-content">

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            <span class="box-title">29/6: Plasmid purified pSB1C3-<I>OBP2A</I> is sent for sequencing  </span>
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                <div id="nmcollapsiblew3-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> Sample 5 of the plasmid purified pSB1C3-<i>OBP2A</i> that was done on the 24/6 is sent for sequencing to determine if <I>OBP2A</I> has successfully ligated into pSB1C3.   </p>
                  <p><b>Protocols Used:</b> N/A</p>
         <p><b>Results: </b> N/A 

                <p><b>Next Steps:</b> If sequencing comes back positive, then the next step will be to clone <i>OBP2A</i> into the two vectors of the bacterial two hybrid system.  </p>
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      </div>


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            <span class="box-title">30/6: Overnight culture of pSB1C3-<I>OBP2A</I> from  the sequenced colony  </span>
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                <div id="nmcollapsiblew3-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> Sequencing came back positive for pSB1C3-<I>OBP2A</I>. So an overnight culture of pSB1C3-<I>OBP2A</I>  will be done in preparation for plasmid purification.     </p>
                  <p><b>Protocols Used:</b><a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a>  </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> Once the overnight culture has been given ample time to grow, the culture will then undergo a plasmid purification. </p>
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        </div>
        </div>
      </div>


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            <span class="box-title">1/7: Plasmid Purification of pSB1C3-<I>OBP2A</I>  </span>
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                <div id="nmcollapsiblew3-3" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To produce a plasmid purification of the sequenced pSB1C3-<I>OBP2A</I> from the overnight culture set up on the 30/6.   </p>
                  <p><b>Protocols Used:</b><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification </a>  </p>
         <p><b>Results: </b> The plasmid purified pSB1C3-<i>OBP2A</i> produced a concentration of 448.83 ng/ul.  </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation is to use the plasmid purification of pSB1C3-<i>OBP2A</i> as the template for PCR using primers that are designed to separate the gene for use in the two hybrid system.  </p></div>
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        </div>
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<!-- WEEK 4 -->
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        <div class="labtitle">Week Beginning 12/7/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Ran PCR of <I>OBP2A</I> using primers for insertion of <I>OBP2A</I> into pUT18 and pT25. </p>
         </div>
         <div id="nmcollapsiblew4" class="collapse week-content">

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       <div class="border-day">
        <div class="box">
            <span class="box-title">17/7: PCR of <I>OBP2A</I> for insertion into bacterial two hybrid vectors: pUT18 and pT25 </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew4-1"></span>
                <div id="nmcollapsiblew4-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a PCR of <I>OBP2A</I> for its insertion into the bacterial two hybrid vectors; pUT18 and pT25. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b>The next stage of experimentation will be to gel extract and then subsequently restrict the amplified gene fragments of <i>OBP2A</i>.  </p>
                </div>
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        </div>
      </div>
             

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  </div>
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<!-- WEEK 5 -->
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     <div class="box">
        <div class="labtitle">Week Beginning 20/7/2015</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Attempted insertion of <I>OBP2A</I> into the vectors for the bacterial two hybrid system; pUT18 and pT25.  </p>
         </div>
         <div id="nmcollapsiblew5" class="collapse week-content">

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       <div class="border-day">
        <div class="box">
            <span class="box-title">20/7: Gel Extraction and Restriction Digest of <I>OBP2A</I> from the PCR performed on 17/7  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5-1"></span>
                <div id="nmcollapsiblew5-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a gel extraction and then subsequently a restriction of <I>OBP2A</I> from the PCR produced on the 17/7 using primers designed for the insertion of <i>OBP2A</i> into the vectors for the bacterial two hybrid system, pUT18 and pT25.   </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digest </a></p>
         <p><b>Results: </b>N/A</p> 
                  <p><b>Next Steps:</b> If the gel extraction and subsequent restriction is successful, then the next step will be to ligate the restricted <i>OBP2A</i> into the bacterial two hybrid vector, pUT18 and pT25.   </p>
                </div>
        </div>
        </div>
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        <div class="box">
            <span class="box-title">21/7: Ligation and Transformation of <I>OBP2A</I> into pUT18 and pT25.     </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5-2"></span>
                <div id="nmcollapsiblew5-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a ligation and  then subsequently a transformation  of <I>OBP2A</I> into the vectors for the bacterial two hybrid system, pUT18 and pT25. Whilst using JM110 <I>E.coli</I> cells as a chassis for transformation. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation </a></p>
         <p><b>Results:  N/A </b></p> 
                  <p><b>Next Steps:</b> The results from the transformation will be seen tomorrow. If successful and positive colonies form on the antibiotic plates, then the next step will be to grow overnight colonies in preparation for purification and sequencing.    </p>
                </div>
        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">22/7: Re-ligation and Transformation of <I>OBP2A</I> into pUT18 and PT25.
  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5-3"></span>
                <div id="nmcollapsiblew5-3" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a re-ligation and transformation of <i>OBP2A</i> into pUT18 and pT25 - since the transformation performed on the 21/7 failed. 
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligation</a> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation </a></p>
         <p><b>Results: N/A </b></p> 
                  <p><b>Next Steps:</b> If transformation is successful and positive colonies form on the antibiotic plates, then the next step will be to grow overnight colonies in preparation for purification and sequencing.   </p>
                </div>
        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">23/7: Production of overnight cultures of pUT18-OBP2A. </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5-4"></span>
                <div id="nmcollapsiblew5-4" class="collapse box-content">
                  <p><b>Aim of Experiment:</b>  To produce overnight cultures of pUT18-<i>OBP2A</i>. Unfortunately no positive colonies were produced from the transformation of <i>OBP2A</i> into pT25.    </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Culture </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> Once the cells have been given ample time to grow, they will undergo plasmid purification for subsequent sequencing.  </p>
                </div>
        </div>
        </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">24/7: Plasmid Purification and subsequent Presequence digest of pUT18-<I>OBP2A</I></span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew5-5"></span>
                <div id="nmcollapsiblew5-5" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a plasmid purification of  the overnight cultures of pUT18-<i>OBP2A</i> and then a subsequent presequence digest.  
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification</a> <a data-toggle="modal" data-target="#presqunce digest-modal" class="journal-protocol">Restriction Digest</a></p>
         <p><b>Results:</b> <a data-toggle="modal" data-target="#nm-figure5-modal" class="journal-protocol"> Figure 5 </a></p> 
                  <p><b>Next Steps:</b> If sequencing comes back positive for pUT18-<I>OBP2A</I> then this can now be implemented into the bacterial two hybrid system. Another attempt at cloning <I>OBP2A</I> into pT25 will be done next week.  </p>     </div>    
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        </div>
        </div>
      </div>


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  </div>
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<!-- WEEK 6 -->
  <div class="row">
    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 27/7/2015</div>
         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6"></span>
          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Successful insertion of the two subunits of <I>OBP2A</I> into the vectors pUT18 and pT25.   </p>
         </div>
         <div id="nmcollapsiblew6" class="collapse week-content">
<div class="row">
       <div class="border-day">
        <div class="box">
            <span class="box-title">27/7: PCR  and Gel extraction of <I>OBP2A</I> for Preparation of Insertion into  pT25.   </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6-1"></span>
                <div id="nmcollapsiblew6-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a PCR and subsequent gel extraction of <i>OBP2A</i> for its insertion into pT25.  </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a> </p>
         <p><b>Results: </b><a data-toggle="modal" data-target="#nm-figure6-modal" class="journal-protocol"> Figure 6 </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to restrict <I>OBP2A</I> for subsequent ligation into pT25.    </p>
                </div>
        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">28/7: Restriction Digest of <I>OBP2A</I> for pT25 and Positive Sequencing of pUT18-<I>OBP2A</I>     </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6-2"></span>
                <div id="nmcollapsiblew6-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a restriction digest of <I>OBP2A</I> for subsequent ligation into PT25. Also, sequencing came back positive for pUT18-<I>OBP2A</I>, so pUT18-<I>OBP2A</I> can now be implemented in the bacterial two hybrid system.  </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#restrictiondigest-modal" class="journal-protocol">Restriction Digests</a> </p>
         <p><b>Results: </b>N/A </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to ligate <I>OBP2A</I> into the pT25 vector.    </p>
                </div>
        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">30/7: Ligation and Transformation of <I>OBP2A</I> into pT25.    </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6-3"></span>
                <div id="nmcollapsiblew6-3" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a ligation and transformation of <I>OBP2A</I> into the bacterial two hybrid vector pT25, so that the interaction between the two constituents of <i>OBP2A</i> can be observed using the bacterial two hybrid system.   </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligation</a><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </p>
         <p><b>Results: N/A</p> 
                  <p><b>Next Steps:</b> If the transformation is successful and positive colonies form on the antibiotic plates, then the next step will be to grow overnight cultures of those colonies in preparation for purification and sequencing.   </p>
                </div>
        </div>
        </div>
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        <div class="box">
            <span class="box-title">31/7: Overnight Cultures of pT25-<I>OBP2A</I> from Positive Colonies. </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6-4"></span>
                <div id="nmcollapsiblew6-4" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To produce overnight cultures of pT25-<i>OBP2A</i> from the transformation done on the 30/7. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Culture </a> 
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> Once the cells have grown overnight they will undergo plasmid purification for subsequent sequencing.  </p>
                </div>
        </div>
        </div>
      </div>
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        <div class="box">
            <span class="box-title">1/8: Plasmid Purification and Subsequent Presequence Restriction Digest of pT25-<I>OBP2A</I> </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew6-5"></span>
                <div id="nmcollapsiblew6-5" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a plasmid purification of pT25-<i>OBP2A</i>  from the overnight cultures done on the 31/7 and then a subsequent presequence digest.    </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid Purification</a><a data-toggle="modal" data-target="#pre-sequence digest-modal" class="journal-protocol"> Restriction Digest</a>
         <p><b>Results: </b> <a data-toggle="modal" data-target="#nm-figure7-modal" class="journal-protocol"> Figure 7 </a></p> 
                  <p><b>Next Steps:</b> If sequencing comes back positive for pT25-<I>OBP2A</I> then this can now be implemented into the bacterial two hybrid system. The results from the sequencing should arrive early on the 3/8.</div>     
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<!-- WEEK 7 -->
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     <div class="box">
        <div class="labtitle">Week Beginning 3/8/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content">Positive sequencing of pT25-<i>OBP2A</i>, subsequent transformation into the bacterial two hybrid system and plating onto MacConkey agar.    </p>
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            <span class="box-title">3/8: Transformation of pUT18-<I>OBP2A</I> into BTH101 <I>E.coli</I> strain.    </span>
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                  <p><b>Aim of Experiment:</b> To perform a transformation of pUT18-<i>OBP2A</i> into BTH101, so that a subsequent transformation of pT25-<i>OBP2A</I> into the same chassis can occur as both plasmids are required to reside in the same chassis so that their interaction can be observed.    </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> If the transformation is successful, then the next stage of experimentation will be to re-transform this culture of cells using the pT25-<I>OBP2A</I> plasmid.      </p>
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            <span class="box-title">4/8:Day culture and transformation of BTH101 containing pUT18-<I>OBP2A</I> with pT25<I>OBP2A</I>   </span>
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                  <p><b>Aim of Experiment:</b> To perform a day culture and then a subsequent transformation of the BTH101 chassis containing  pUT18-<i>OBP2A</i> from the transformation performed on 3/8.  </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> If the transformation is successful then the next stage of experimentation will be to observe the  protein interaction of the two subunits of <I>OBP2A</I> using MacConkey agar plates.     </p>
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            <span class="box-title">5/8:Day culture and plating of positive colonies from transformation onto MacConkey agar.  </span>
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                  <p><b>Aim of Experiment:</b> To perform a day culture and then subsequent plating of positive colonies from the transformation performed on the 4/8. Positive colonies from this transformation contain both vectors of the bacterial two hybrid system along with the two subunits of <i>OBP2A</i> and so now the interaction can be observed. The day culture will be plated onto MacConkey agar which once the cells have been given enough time to grow will give a visual demonstration of the  protein interaction through a color change.      </p>
                  <p><b>Protocols Used:</b> </a><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </p>
         <p><b>Results: </b>N/A </a></p> 
                  <p><b>Next Steps:</b> It has been decided that whilst the <I>E.coli</I> strain BTH101 that is being used in this experiment is useful for visualizing protein interaction, it is very unstable. A better <I>E.coli</I> strain called MG1655(Δcya) will be used instead as it is much more stable. This means that tomorrow overnights will be prepared of MG1655(Δcya)for transformation with pUT18-<i>OBP2A</i> and then subsequently pT25-<i>OBP2A</i>.   </p>
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            <span class="box-title">6/8: Overnight cultures of MG1655(Δcya)</span>
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                  <p><b>Aim of Experiment:</b> To perform an overnight culture of MG1655(Δcya)<I>E.coli</I> in preparation for transformation with pUT18-<I>OBP2A</I> and then pT25-<I>OBP2A</I>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Culture </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> Once the overnight culture has been give sufficient time to grow these cells will then be transformed with pUT18-<I>OBP2A</I> and then subsequently re-transformed with pT25-<I>OBP2A</I>.   </p>
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            <span class="box-title">7/8: Results of MacConkey agar growth and transformation of  MG1655(Δcya)with pUT18-<I>OBP2A</I> </span>
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                <div id="nmcollapsiblew7-5" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a transformation of MG1655(Δcya)with pUT18-<I>OBP2A</I>. Also the results of plating of our bacterial two hybrid interaction onto MacConkey agar has produced promising results as the colonies appear to be taking up a small amount of red dye - an indicator of protein interaction. This suggests that the two separated parts of <I>OBP2A</I> may in fact be interacting in vivo. This interaction will need to be quantified through the use of the β-galactosidase assay. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> If the transformation of MG1655(Δcya) with pUT18-<I>OBP2A</I> is successful then the next step of experimentation will be re-transform positive colonies with pT25-<I>OBP2A</I>.  </p>
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            <span class="box-title">8/8: Transformation of MG1655(Δcya) containing pUT18-<I>OBP2A</I> with pT25-<i>OBP2A</i>  </span>
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                <div id="nmcollapsiblew7-6" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a transformation of MG1655(Δcya) containing pUT18-<I>OBP2A</I> with pT25-<i>OBP2A</i>. Positive colonies were produced from the transformation done on the 7/7 and so as a result a day culture was produced and subsequently re-transformed with pT25-<i>OBP2A</i>. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> If the transformation of MG1655(Δcya) with pT25-<I>OBP2A</I> is successful then the next step of experimentation will be to undertake the β-galactosidase assay  </p></div>
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        </div>
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      </div>

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  </div>
</div>



<!-- WEEK 8 -->
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        <div class="labtitle">Week Beginning 10/8/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> First attempt at determination of the protein interaction of the <I>OBP2A</i> subunits using the β-galactosidase assay.   </p>
         </div>
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            <span class="box-title">10/8: Plating of Controls in preparation of the β-galactosidase assay   </span>
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                <div id="nmcollapsiblew8-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To plate control interactions that are used in the β-galactosidase assay to aid in the interpretation of the results form the assay. This first assay attempt will consist of one positive control, one negative control and one regulatory control.       </p>
                  <p><b>Protocols Used: N/A </b></p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to perform overnight cultures from positive colonies in preparation for the β-galactosidase assay.      </p>
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        </div>
        </div>
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        <div class="box">
            <span class="box-title">11/8: Overnight cultures of controls and <i>OBP2A</i> in preparation for the β-galactosidase assay.  </span>
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                <div id="nmcollapsiblew8-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To prepare overnight cultures of positive colonies of MG1655(Δcya) containing pUT18-<I>OBP2A</I> and pT25-<I>OBP2A</I>, along with the controls in preparation for the  β-galactosidase assay. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> Once the cells have been given sufficient time to grow then the next stage of experimentation will be to inoculate those colonies into fresh liquid broth until they reach an O.D of 0.4 before being pellet and frozen for the assay.    </p>
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        </div>
        </div>
      </div>
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        <div class="box">
            <span class="box-title"> 12/8: Inoculation and preparation of cultures for the  β-galactosidase assay.    </span>
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                <div id="nmcollapsiblew8-3" class="collapse box-content">
                  <p><b>Aim of Experiment:</b>  To prepare cultures for use in the  β-galactosidase assay, this requires a small subsample of each overnight to be inoculated into fresh LB, two done for each colony. Then left to grow until the O.D of each culture is between 0.3-0.5. Once an O.D of 0.3-0.5 has been reached then 3 replicas of 1 ml samples are taken from each culture before being spun down, the supernatant then subsequently removed and the pellet frozen.      </p>
                  <p><b>Protocols Used:</b> </a><a data-toggle="modal" data-target="#ap-modal" class="journal-protocol"> Assay Preparation</a> </p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> These samples are now ready for the β-galactosidase assay.   </p>
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        </div>
        </div>
      </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">13/8: First attempt at the determination of the protein interaction of the two <I>OBP2A</i> subunits using the β-galactosidase assay </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew8-4"></span>
                <div id="nmcollapsiblew8-4" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To determine the millers activity of the two interacting subunits of <I>OBP2A</I> in vivo as a basis of the level of interaction between them. This was done using the β-galactosidase assay. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#bga-modal" class="journal-protocol">β-galactosidase assay</a> 
         <p><b>Results: </b> <a data-toggle="modal" data-target="#nm-figure8-modal" class="journal-protocol"> Figure 8 </a></p> 
                  <p><b>Next Steps:</b> As it was the first attempt at a β-galactosidase assay, subsequent assays will be performed utilizing more controls. </p>
                </div>
        </div>
        </div>
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        <div class="box">
            <span class="box-title">14/8: Preparation of overnight cultures of controls and samples in preparation of performing a second β-galactosidase assay    </span>
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                <div id="nmcollapsiblew8-5" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform overnight cultures of the controls and samples that will be necessary for doing this second β-galactosidase assay. This assay will include two positive controls which are known to have a high level of interaction - these are NarG-NarJ and Zip positive controls. There will also be two negative controls, an empty MG1655(Δcya) chassis and pUT18-<i>OBP2A</i> with NarJ. Finally there is one regulatory control: which contains two empty vectors of pUT18 and pT25.     </p>
                  <p><b>Protocols Used:</b><a data-toggle="modal" data-target="#onc-modal" class="journal-protocol">Overnight Cultures </a></p>
         <p><b>Results: </b>N/A </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to prepare samples for use in the β-galactosidase assay.  </p></div>
                </div>
        </div>
        </div>
      </div>

    </div>
  </div>
</div>

<!-- WEEK 9 -->
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    <div class="border-week">
     <div class="box">
        <div class="labtitle">Week Beginning 17/8/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Second attempt at determining the protein interactions of the two <I>OBP2A</i> subunits using the β-galactosidase assay and subsequent cloning of <i>OBP2A</i> into the bacterial two hybrid vectors in switched order.  </p>
         </div>
         <div id="nmcollapsiblew9" class="collapse week-content">
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       <div class="border-day">
        <div class="box">
        <span class="box-title">17/8: Preparation of the controls and samples for use in the second attempt of the β-galactosidase assay  </span>
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                <div id="nmcollapsiblew9-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To prepare cultures for use in the  β-galactosidase assay, this requires a small subsample of each overnight to be inoculated into fresh LB, two done for each colony. Then left to grow until the O.D of each culture is between 0.3-0.5. Once an O.D of 0.3-0.5 has been reached then 3 replicas of 1 ml samples are taken from each culture before being spun down, the supernatant then subsequently removed and the pellet frozen. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#bga-modal" class="journal-protocol">β-galactosidase assay  </a> </p>
         <p><b>Results: </b>N/A </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to perform the second β-galactosidase assay on the interaction between the two subunits of <i>OBP2A</i>      </p>
                </div>
        </div>
        </div>
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       <div class="border-day">
        <div class="box">
            <span class="box-title">18/8: Second attempt at determination of the protein interactions of the <I>OBP2A</i> subunits using the β-galactosidase assay  </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew9-2"></span>
                <div id="nmcollapsiblew9-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To perform a second attempt at the β-galactosidase assay that will aid in the determination of the level of interaction that the two separated subunits of <i>OBP2A</i> are inflicting on one another. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#bga-modal" class="journal-protocol">β-Galactosidase assay  </a> </p>
         <p><b>Results: </b><a data-toggle="modal" data-target="#nm-figure11-modal " class="journal-protocol"> Figure 10 </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to switch the positioning of the two subunits of <i>OBP2A</i> in pUT18 and pT25 by having the smaller subunit in pUT18 and the large subunit in pT25. Along with this another β-galactosidase assay will be performed however this time it will be done under anaerobic conditions to see if that has any appreciable effect on the interaction of the two subunits of <i>OBP2A</i>.      </p>
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        <div class="box">
            <span class="box-title"> 20/8: PCR of <i>OBP2A</i> in preparation of pUT18 and pT25 insertion   </span>
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                <div id="nmcollapsiblew9-3" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To  Perform a PCR of <i>OBP2A</i> using primers for pU1T18 and pT25 insertion which have been designed to place the smaller <i>OBP2A</i>subunit into pUT18 and the larger subunit into pT25. </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a> 
         <p><b>Results: </b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> N/A </a></p> 
                  <p><b>Next Steps:</b> If PCR is successful then the next stage of experimentation will be to perform a gel extraction and restriction in perpetration for ligation and transformation of <i>OBP2A</i> into pUT18 and pT25.   </p>
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        <div class="box">
            <span class="box-title">21/8: Second attempt at a PCR of <i>OBP2A</i> in preparation of pUT18 and pT25 insertion   </span>
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                <div id="nmcollapsiblew9-4" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> The PCR performed on the 20/8 failed as no bands were found to be present on the gel during gel viewing. As a result of this a subsequent PCR was done today.      </p>
                  <p><b>Protocols Used:</b> </a><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a> </p>
         <p><b>Results: </b> N/A  </a></p> 
                  <p><b>Next Steps:</b>  If PCR is successful then the next stage in experimentation will be to perform a gel extraction and restriction in perpetration for ligation and transformation of <i>OBP2A</i> in pUT18 and pT25.  </p>
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        </div>
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            <span class="box-title">22/8: Second attempt at PCR of <i>OBP2A</i> for insertion into pUT18 and pT25 failed </span>
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                <div id="nmcollapsiblew9-5" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> A second attempt at preparing <i>OBP2A</i> for insertion into the two vectors of the bacterial two hybrid system(pUT18 and pT25) has failed! As a result this will be left until next week, so that the rest of the week can be dedicated to preparing and undertaking  another β-galactosidase assay, that will be done under anaerobic conditions.  </p>
                  <p><b>Protocols Used:</b> N/A </a></p>
         <p><b>Results: </b> N/A </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation is to prepare for a anaerobic β-galactosidase assay of <i>OBP2A</i> in the bacterial two hybrid vectors; pUT18 and pT25.  </p></div>
                  </div>
        </div>
        </div>
      </div>
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  </div>
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  <!-- WEEK 10-->
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     <div class="box">
        <div class="labtitle">Week Beginning 24/8/2015</div>
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          <div class="box-content">
          <p class="journal-summary-heading">Summary</p>
            

         <p class="journal-content"> Performed an anaerobic β-galactosidase assay in order to determine the level of interaction between the two subunits of <i>OBP2A</i>.   </p>
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        <div class="box">
            <span class="box-title">24/8: Preparation of samples for the anaerobic β-galactosidase assay of <i>OBP2A</i> in pUT18 and pT25  </span>
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                <div id="nmcollapsiblew10-1" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> To prepare the  samples and controls of <i>OBP2A</i> for the anaerobic β-galactosidase assay, so they can be frozen overnight in preparation for the assay itself.    </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#ap-modal" class="journal-protocol">Assay Preparation</a> </p>
         <p><b>Results: </b> </a></p> 
                  <p><b>Next Steps:</b> The next stage of experimentation will be to perform the β-galactosidase assay itself. 
               </div>
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        </div>
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        <div class="box">
            <span class="box-title"> 25/8: Performed an anaerobic β-galactosidase assay in order to determine the level of interaction of <i>OBP2A</i> in the bacterial two hybrid vectors: pUT18 and pT25.   </span>
              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#nmcollapsiblew10-2"></span>
                <div id="nmcollapsiblew10-2" class="collapse box-content">
                  <p><b>Aim of Experiment:</b> Performed an anaerobic β-galactosidase assay of <i>OBP2A</i> in the bacterial two hybrid vectors: pUT18 and pT25. This is to determine the level of interaction that the smaller subunit of <i>OBP2A</i> has with its larger subunit in vivo. This particular assay has two positive controls, two negative controls and one regulatory control.  </p>
                  <p><b>Protocols Used:</b> <a data-toggle="modal" data-target="#bga-modal" class="journal-protocol"> β-galactosidase assay </a> </p>
         <p><b>Results: </b><a data-toggle="modal" data-target="#nm-figure12-modal" class="journal-protocol"> Figure 12 </a></p> 
                  <p><b>Next Steps:</b> The results have shown that the two subunits of <i>OBP2A</i> in fact don't seem to be interacting in vivo - a possible reason for this might be aggregation of the two subunits preventing interaction. The  next stage of experimentation will be to test out the interaction when the subunit are switch in relation to heir positions on the vectors: pUT18 and pT25.    </p>
                </div>
        </div>
        </div>
      </div>
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