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| − | <h1>Under Construction</h1>
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| − | <!--<IMG SRC="https://static.igem.org/mediawiki/2015/e/ea/Viable_count_E.coli_DH5alpha_checking_quality_team_Birkbeck_iGEM_2015.jpg">-->
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| − | <!--<p>The growth kinetics of E. coli DH5α had to be established if experiments involving infection with λ-bacteriophage and characterizing a potential signal in living cells. It was important for the study to also quantify the number of viable cells with relation to the optical density of the culture.</p>
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| − | <center><IMG SRC="https://static.igem.org/mediawiki/2015/5/58/Growth_Curve_50_mL_600_nm_team_birkbeck_iGEM_2015.jpg"></center>
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| − | <p><b><u>Fig. 1: Growth Curve of <i>E. coli</i> DH5α Strains Following Culture Optical Density of 600 nm</u></b>.</p>
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| − | <br>
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| − | <p>Growth kinetics was initially studied using 50 mL cultures. <b>Fig. 1</b> shows the growth kinetics of <i>E. coli</i> DH5α & derivative strains containing plasmids from the <a href="https://2015.igem.org/Team:Birkbeck/InterLab_Study">InterLab</a> study. The growth curve shows that the <i>E. coli</i> strain that contains the P1-<i>gfp</i> expression device grows at a slower rate than the other strains investigated. At 220 minutes the <i>E. coli</i> DH5α P1 strain has a significantly lower OD<sub>600</sub> than the <i>E. coli</i> DH5α (P=0.023). <i>E. coli</i> DH5α remains significantly higher in OD<sub>600</sub> than <i>E. coli</i> DH5α with the P1-<i>gfp</i> expression device (P=<0.001)<!---->. The only difference between the <i>E. coli</i> DH5α & <i>E. coli</i> DH5α positive control device is observed at 280 minutes into the growth curve (P=0.016) where the positive control has a higher OD<sub>600</sub>. The multiple comparison table showing P values can be viewed in <a href="https://2015.igem.org/Team:Birkbeck/Results/Table_S1"><b>Table S1</b>.</a></p>.
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| − | <center><img src="https://static.igem.org/mediawiki/2015/a/a8/Growth_Curve_%28395_nm%29_50_mL_Cultures_Team_birkbeck_iGEM_2015.jpg "></center>
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| − | <p><b><u>Fig. 2: Growth Curve of <i>E. coli</i> DH5α Strains Following Culture Optical Density of 395 nm</u></b>.</p>
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| − | <br>
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| − | <p>In order to investigate if there could be a point in the <i>E. coli</i> DH5α growth curve in which a signal from the GFP could be detected by absorbance, the growth curves were also conducted using the major absorption peak of GFP (wavelength 395 nm)<!--(REF!)-->. The growth curve data for the culture optical density is displayed in <b>Fig. 2</b>.</P>
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| − | <p>When comparing the data point of <i>E. coli</i> DH5α strains, there appears to be a significant increase in culture OD<sub>395</sub> in the <i>E. coli</i> cells with the P1-<i>gfp</i> expression device (P<0.001). This apparent signal is only present between 60-100 minutes of growth. When comparing the positive control & <i>E. coli</i> DH5α1, there is no significant difference between the data sets at 60 or 100 minutes (P=1 for both time points). A small potential signal is observed from the oositive control <i>gfp</i> expression device at 280 minutes (P=0.012) & 300 minutes (P=0.006). This significance is lost after 300 minutes (P=0.262). See <a href="https://2015.igem.org/Team:Birkbeck/Results/Table_S2"><b>Table S2</b></a> for more details. In order to verify these results & to test for the feasibility of scaling down to a 96-well microtitre plate assay, a 10 hour growth curve was conducted in a 96-well microtitre plate (see <b>Fig. 6-8</b>).</p>
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| − | <!--Multiple Comparison table link for OD 395 nm (table S2); https://2015.igem.org/Team:Birkbeck/Results/Table_S2. issue sorted, this page is good to go!.-->
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| − | <!--For sorting out code to make it look decent on our wiki; https://2015.igem.org/Team:Birkbeck/Results/adjusting_table_code ; use this link. its a spoof web page.-->
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| − | <br> | + | <head> |
| − | <center><img src="https://static.igem.org/mediawiki/2015/d/d7/Viable_count_1_hour_%28dh5_alpha%29_team_birkbeck_iGEM_2015.jpg"></center>
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| − | <p><b><u>Fig. 3: Viable Count of <i>E. coli</i> DH5α After 60 mins</u></b>.</p>
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| − | <br>
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| − | <p>In order to assess how many viable cells correspond to different optical densities, a viable count was conducted on <i>E. coli</i> DH5α1 at 60 minutes (<b>Fig. 3</b> & <b>Table 1</b>), 175 minutes (<b>Fig. 5</b> & <b>Table 2</b>) & 225 minutes (data not shown due to high level of contamination).</p>
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| − | <p>Considering the OD<sub>600</sub> of the <i>E. coli</i> DH5α1 cultures at 60 mins, triplicate cultures were OD<sub>600</sub> = 0.029, 0.01 & 0.025<!--0.021 mean-->. The viable count of each of the cultures gave a mean of 2.57×10<sup>6</sup> cfu/mL (<b>Table 1</b>). It can therefore be concluded that an <i>E. coli</i> DH5α culture at an OD<sub>600</sub> = 0.021 corresponds to approximately 2.57×10<sup>6</sup> cfu/mL.</p>
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| − | <caption class="title" style="width:522px">Descriptive Statistics of 1 hour Viable Count of <i>E. coli</i> DH5α.<span class="details"></span></caption> | + | |
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| − | <td class="columnLabels dataAreaLeft vCC role3">N</td>
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| − | <td class="columnLabels vCC role3">Minimum</td>
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| − | <td class="columnLabels vCC role3">Maximum</td>
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| − | <td class="columnLabels vCC role3">Mean</td>
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| − | <td class="columnLabels vCC role3">Std. Deviation</td>
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| − | <td class="rowLabels dataAreaTop role3">Viable Count</td>
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| − | <td class=" e dataAreaTop dataAreaLeft vCC">9</td>
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| − | <td class=" e dataAreaTop vCC">1600000</td>
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| − | <td class=" e dataAreaTop vCC">3750000</td>
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| − | <td class=" e dataAreaTop vCC">2566666.67</td>
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| − | <td class=" e dataAreaTop vCC">627495.020</td>
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| − | <td class="rowLabels hCR role3">Valid N (listwise)</td>
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| − | <td class=" o dataAreaLeft hCR vCC">9</td>
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| − | </div></body></center>
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| − | <!--End of descriptive Stats table-->
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| − | <p><b><u>Table 1: Descriptive Statistics of 60 minutes Viable Count of <i>E. coli</i> DH5α.</u></b>.</p>
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| − | <br> | + | |
| − | <center><img src="https://static.igem.org/mediawiki/2015/e/e6/Viable_count_of_DH5_alpha_after_175_mins_team_birkbeck_iGEM_2015.jpg"></center>
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| − | <p><b><u>Fig. 4: Viable Count of <i>E. coli</i> DH5α After 175 mins</u></b>.</p>
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| − | <br>
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| − | <p>At 175 minutes into growth, the <i>E. coli</i> DH5α1 cultures had OD<sub>600</sub> of; 0.255, 0.216 & 0.262<!--mean = 0.244-->. The viable count of each of the cultures gave a mean of 1.33×10<sup>8</sup> cfu/mL (<b>Table 2</b>). It can therefore be concluded that an <i>E. coli</i> DH5α culture at an OD<sub>600</sub> = 0.244 corresponds to approximately 1.33×10<sup>8</sup> cfu/mL. Considering the previous OD<sub>600</sub> (0.021), there is approximately a 10-fold increase in OD<sub>600</sub> which corresponds to nearly a 100-fold increase in viable cells.</p>
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| − | <!--Begining of the descriptive statistics table for 175 minutes of growth--> </p>
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| − | </style>
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| − | </div>
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| − | <center><div class="itemBody">
| + | |
| − | <table class="Default PivotTable" data-lookName="Default" data-rowLabelWidth="1" data-colLabelHeight="1" data-NumRows="2" data-NumCols="5" data-rowLabels="Corner" data-decimal-char="." data-col-labels="N;Minimum;Maximum;Mean;Std. Deviation" data-row-labels="Viable.Count;Valid N (listwise)" >
| + | |
| − | <caption class="title" style="width:547px"><span class="details">Descriptive Statistics of 175 Minutes Viable Count of <i>E. coli</i> DH5α.</span></caption>
| + | |
| − | <tbody class="PivotBody defaultLayer innerFrameBd"> | + | |
| − | <tr>
| + | |
| − | <td class="cornerLabels"> </td>
| + | |
| − | <td class="columnLabels dataAreaLeft vCC role3">N</td>
| + | |
| − | <td class="columnLabels vCC role3">Minimum</td>
| + | |
| − | <td class="columnLabels vCC role3">Maximum</td>
| + | |
| − | <td class="columnLabels vCC role3">Mean</td>
| + | |
| − | <td class="columnLabels vCC role3">Std. Deviation</td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td class="rowLabels dataAreaTop role3">Viable Count</td>
| + | |
| − | <td class=" e dataAreaTop dataAreaLeft vCC">9</td>
| + | |
| − | <td class=" e dataAreaTop vCC">90000000</td>
| + | |
| − | <td class=" e dataAreaTop vCC">175000000</td>
| + | |
| − | <td class=" e dataAreaTop vCC">133333333.33</td>
| + | |
| − | <td class=" e dataAreaTop vCC">29154759.474</td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td class="rowLabels hCR role3">Valid N (listwise)</td>
| + | |
| − | <td class=" o dataAreaLeft hCR vCC">9</td>
| + | |
| − | <td class=" o hCR vCC"> </td>
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| − | <td class=" o hCR vCC"> </td>
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| − | </tr>
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| − | </tr>
| + | |
| − | </tbody>
| + | |
| − | </table>
| + | |
| − | </div></body></center>
| + | |
| − | <!--End of the descriptive statistics table for 175 minutes of growth-->
| + | |
| − | <p><b><u>Table 2: Descriptive Statistics of 175 minutes Viable Count of <i>E. coli</i> DH5α.</u></b></p>
| + | |
| − | <br>
| + | |
| − | <center><IMG SRC="https://static.igem.org/mediawiki/2015/8/8a/Growth_curve_601_nm_96-microtitre_Team_birkbeck_iGEM_2015_data.jpg "></center>
| + | |
| − | <p><b><u>Fig. 5: Growth Curves of Different Strains of <i>E. coli</i> DH5α Following Culture Optical Density at 601 nm</u></b>.</p> | + | |
| − | <br>
| + | |
| − | <p><a href="https://2015.igem.org/Team:Birkbeck/table_S3">Table S3</a>, <a href="https://2015.igem.org/Team:Birkbeck/table_S5"></a>Table S5</p> & <a href="https://2015.igem.org/Team:Birkbeck/table_S6">Table S6</a>
| + | |
| − | <!--Table S3 (0-200 mins); https://2015.igem.org/Team:Birkbeck/table_S3, (change to S4!!!)Tables S5 (220-420); https://2015.igem.org/Team:Birkbeck/table_S5, Table S6 (440-580 minutes); https://2015.igem.org/Team:Birkbeck/table_S6 -->
| + | |
| − | <br>
| + | |
| − | <center><IMG SRC="https://static.igem.org/mediawiki/2015/d/d0/Growth_curve_501_nm_microtitre_data_Team_birkbeck_iGEM_2015.jpg"></center>
| + | |
| − | <p><b><u>Fig. 6: Growth Curves of Different Strains of <i>E. coli</i> DH5α Following Culture Optical Density at 501 nm</u></b>.</p>
| + | |
| − | <br>
| + | |
| − | <p><a href="https://2015.igem.org/Team:Birkbeck/table_S7">Table S7</a>, <a href="https://2015.igem.org/Team:Birkbeck/table_S8">Table S8</a> & <a href="https://2015.igem.org/Team:Birkbeck/table_S9">Table S9</a></p>
| + | |
| − | <!--Table S7 (0 mins - 200 mins); https://2015.igem.org/Team:Birkbeck/table_S7 , Table S8 (240-420 minutes); https://2015.igem.org/Team:Birkbeck/table_S8 Table S9 (440-580 mins); https://2015.igem.org/Team:Birkbeck/table_S9-->
| + | |
| − | <br>
| + | |
| − | <center><img src="https://static.igem.org/mediawiki/2015/4/4a/Growth_curve_475_nm_microtitre_team_birkbeck_iGEM_2015.jpg"></center>
| + | |
| − | <p><b><u>Fig. 7: Growth Curves of Different Strains of <i>E. coli</i> DH5α Following Culture Optical Density at 475 nm</u></b>.</p>
| + | |
| − | <br>
| + | |
| − | <p><a href="https://2015.igem.org/Team:Birkbeck/Table_S10"></a>Table S10</p>, <a href="https://2015.igem.org/Team:Birkbeck/table_S11">Table S11</a> & <a href="https://2015.igem.org/Team:Birkbeck/table_S12">Table S12</a>
| + | |
| − | <!--Table S10 (0 mins-200 mins); https://2015.igem.org/Team:Birkbeck/Table_S10 Table S11 (220-420); https://2015.igem.org/Team:Birkbeck/table_S11 Table S12 (440-580 minutes); https://2015.igem.org/Team:Birkbeck/table_S12-->
| + | |
| − | <br> | + | |
| − | <center><img src="https://static.igem.org/mediawiki/2015/8/8b/Growth_curve_395_microtitre_team_birkbeck_iGEM_2015.jpg "></center>
| + | |
| − | <p><b><u>Fig. 8: Growth Curves of Different Strains of <i>E. coli</i> DH5α Following Culture Optical Density at 395 nm</u></b>.</p>
| + | |
| − | <br>
| + | |
| − | <p><a href="https://2015.igem.org/Team:Birkbeck/Table_S13">Table S13</a>, <a href="https://2015.igem.org/Team:Birkbeck/table_S14">Table S14</a> & <a href="https://2015.igem.org/Team:Birkbeck/table_S15">Table S15</a></p>
| + | |
| − | <!--Table S13 (0-200 minutes); https://2015.igem.org/Team:Birkbeck/Table_S13 Table S14 (220-420 mins); https://2015.igem.org/Team:Birkbeck/table_S14 Table S15 (440-580 mins); https://2015.igem.org/Team:Birkbeck/table_S15 (the P1 is not significant at 440 minutes [P=0.083]-->
| + | |
| − | <br>
| + | |
| | | | |
| − | <center><img src="https://static.igem.org/mediawiki/2015/9/9a/Fluorescence_growth_curve_of_multiple_strains_of_E._coli_DH5_alpha_team_Birkbeck_iGEM_2015.jpg "></center>
| |
| − | <p><b><u>Fig. 9: Growth Curves of Different Strains of <i>E. coli</i> DH5α Following Culture Fluorescence</u></b>.</p>
| |
| − | <br>
| |
| − | <p><a href="https://2015.igem.org/Team:Birkbeck/table_S16">Table S16</a>, <a href="https://2015.igem.org/Team:Birkbeck/Table_S17 ">Table S17</a> & <a href="https://2015.igem.org/Team:Birkbeck/table_S18">Table S18</a></p>
| |
| − | <!--For the fluorescence data, the table has to be split into 3 different pages! bit of a ball-ache but it has to be done its just a really big file. I am guessing the same has to be done with the absorption spectra data :( lots more work needs doing on this matter. Table S16 (0-200); https://2015.igem.org/Team:Birkbeck/table_S16 Table S17 (220-420); https://2015.igem.org/Team:Birkbeck/Table_S17 Table S18 (440 - 580 minutes); https://2015.igem.org/Team:Birkbeck/table_S18-->
| |
| − | <br>
| |
| − | <!--
| |
| − | <h2> Project Results</h2>
| |
| | | | |
| − | <p>Here you can describe the results of your project and your future plans. </p> | + | <div id="maincontainer"> |
| | | | |
| − | <h5>What should this page contain?</h5> | + | <h2><b>Results</b></h2> |
| − | <ul>
| + | |
| − | <li> Clearly and objectively describe the results of your work.</li>
| + | |
| − | <li> Future plans for the project </li>
| + | |
| − | <li> Considerations for replicating the experiments </li>
| + | |
| − | </ul> | + | |
| | | | |
| | + | <div class="accordion"> |
| | | | |
| | + | <center> |
| | + | <h3> |
| | + | <a href="#">BioBrick Cloning Results |
| | + | </h3> |
| | + | <p>Click here to see the results from testing the constructs ORF-314, stf gene, tfa construct, P(Cat)-TetR construct, TetR-controlled tfa circuit and cI-Cro circuit. |
| | + | </p> |
| | + | <br> |
| | + | <!-- ############################################################################# |
| | | | |
| | + | REPLACE WITH PICTURE TO REPRESENT THIS SECTION |
| | | | |
| | | | |
| − | <h4> Project Achievements </h4> | + | <img width="800" height="auto" src="https://static.igem.org/mediawiki/2015/f/f9/20150914_122201.jpg" border="0"/> |
| | | | |
| − | <p>You can also include a list of bullet points (and links) of the successes and failures you have had over your summer. It is a quick reference page for the judges to see what you achieved during your summer.</p>
| |
| | | | |
| − | <ul>
| + | #############################################################################--> |
| − | <li>A list of linked bullet points of the successful results during your project</li>
| + | </a> |
| − | <li>A list of linked bullet points of the unsuccessful results during your project. This is about being scientifically honest. If you worked on an area for a long time with no success, tell us so we know where you put your effort.</li>
| + | </h3> |
| − | </ul> | + | |
| | + | </center> |
| | | | |
| | + | <div> |
| | | | |
| | | | |
| − | <h4>Inspiration</h4> | + | <div style="font-size: 16px;"> |
| − | <p>See how other teams presented their results.</p> | + | <p>To confirm the presence and correct size of the BioBricks we are submitting, all samples were run on an agarose gel. The constructs tested were: |
| | <ul> | | <ul> |
| − | <li><a href="https://2014.igem.org/Team:TU_Darmstadt/Results/Pathway">2014 TU Darmstadt </a></li> | + | <li>ORF-314</li> |
| − | <li><a href="https://2014.igem.org/Team:Imperial/Results">2014 Imperial </a></li> | + | <li>the stf gene</li> |
| − | <li><a href="https://2014.igem.org/Team:Paris_Bettencourt/Results">2014 Paris Bettencourt </a></li> | + | <li>the tfa construct</li> |
| − | </ul> | + | <li>the P(Cat)-TetR construct</li> |
| | + | <li>the TetR-controlled tfa circuit</li> |
| | + | <li>the cI-Cro circuit</li></ul></p> |
| | | | |
| | + | <p>All samples were initially digested with two of the four standard iGEM restriction enzymes (<i>XbaI</i>, <i>EcoRI</i>, <i>SpeI</i> and <i>PstI</i>) and expected fragment sizes calculated (Fig 1a and 2a). Success of the cloning was then determined by comparing actual band sizes to the predicted values (Fig 1b and 2b). |
| | + | </div> |
| | + | <br> |
| | + | |
| | + | <p>a. <IMG SRC="https://static.igem.org/mediawiki/2015/e/e8/Birkbeck_150916_LP_gel1_pred.png" height="600px" width="300px"> |
| | + | b. <IMG SRC="https://static.igem.org/mediawiki/2015/1/18/Birkbeck_150916_LP_gel1.jpeg" height="600px" width="350px"></p> |
| | + | <p><b>Fig 1.</b> Comparison of predicted (a) and actual (b) band sizes on gel 1.</p> |
| | + | <br> |
| | + | <p>a. <IMG SRC="https://static.igem.org/mediawiki/2015/6/60/Birkbeck_150916_LP_gel2_pred.png" height="600px" width="300px"> |
| | + | b. <IMG SRC="https://static.igem.org/mediawiki/2015/f/f9/Birkbeck_150916_LP_gel2.jpeg" height="600px" width="350px"></p> |
| | + | <p><b>Fig 2.</b> Comparison of predicted (a) and actual (b) band sizes on gel 2.</p> |
| | + | |
| | + | </div> |
| | + | <br><hr><br> |
| | + | </div> |
| | + | <br> |
| | + | <!--button for Conclusion--> |
| | + | <a href="https://2015.igem.org/Team:Birkbeck/Conclusion"><img width="400px" height="auto" src="https://static.igem.org/mediawiki/2015/7/7d/Birkbeck_phage_infection2.png" border="0"/></a> |
| | + | <p>To find concluding remarks about our project</p> |
| | + | <!--button for Research--> |
| | + | <a href="https://2015.igem.org/Team:Birkbeck/Research"><img width="400px" height="auto" src="https://static.igem.org/mediawiki/2015/c/c7/Rachel-elliot.jpg" border="0"/></a> |
| | + | <p>Back to main Research page</p> |
| | + | |
| | + | </div> |
| | </div> | | </div> |
| | </html> | | </html> |
b. 
b. 
