Difference between revisions of "Team:Tsinghua-A/Demonstrate"

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<h3>★  ALERT! </h3>
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<p>This page is used by the judges to evaluate your team for the <a href="https://2017.igem.org/Judging/Medals">medal criterion</a> or <a href="https://2017.igem.org/Judging/Awards"> award listed above</a>. </p>
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<p> Delete this box in order to be evaluated for this medal criterion and/or award. See more information at <a href="https://2017.igem.org/Judging/Pages_for_Awards"> Instructions for Pages for awards</a>.</p>
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<h1>Demonstrate</h1>
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<h3>Gold Medal Criterion #4</h3>
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Teams that can show their system working under real world conditions are usually good at impressing the judges in iGEM. To achieve gold medal criterion #4, convince the judges that your project works. There are many ways in which your project working could be demonstrated, so there is more than one way to meet this requirement. This gold medal criterion was introduced in 2016, so check our what 2016 teams did to achieve a their gold medals!
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Please see the <a href="https://2017.igem.org/Judging/Medals">2017 Medals Page</a> for more information.
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              <h2 class="heading-section">Discription here</h2>
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    <div id="apDiv4">Killling Ability Test</div>
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    <div id="apDiv5">I. Background</div>
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    <div id="apDiv6">    &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;The ideal killing ability of the warriors includes the two following characters. First, the killing ability of the warriors from the two sides must be roughly equal. Second, the killing ability of the warriors must be adequate——neither too strong nor too weak.</div>
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    <div id="apDiv7">II.Experimental design</div>
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    <div id="apDiv8">&nbsp;&nbsp;&nbsp;&nbsp;To test if the killing ability of the warriors from two sides are the same, we let equal amount of the two kinds of warriors to grow together. We attach different fluorescins to different warriors in order to differentiate them. We compare the killing ability of the warriors by monitoring the ratio of the two kinds of the warriors.<br>
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&nbsp;&nbsp;&nbsp; To test if the killing ability is adequate or not, we do a gradient experiment. We let warriors and beggars from different sides to grow together with different starting ratios. The warriors and beggars from different sides are attached to different fluorescins. We monitor the change of the amount of beggars with time to see if the killing ability is ideal or not. </div>
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    <div id="apDiv9">III Methods</div>
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    <div id="apDiv10">1. Construct our warriors and beggars by co-transformation. (the gene circuits are as follows) *if you want to know the details of how we assemble the parts together, see our protocol “DNA construction” </div>
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    <div id="apDiv11"><img src="circuit.png" width="791" height="492"></div>
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    <div id="apDiv12">Fig 1 gene circuits of warriors and beggars from both sides</div>
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    <div id="apDiv13">2. Pick bacterial clones from the petri plate, then shake it overnight in the LB medium (3ml) with 50μg/ml Ampicilin and 30μg/ml Kanamycin at 37℃. For each combination, 3 clones are picked. The clones are numbered from 1 to 3.  <br>
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      3. Dilute the overnight culture to 1/50 of the original density in fresh LB medium (5ml) containing 50μg/ml Ampicilin and 30μg/ml Kanamycin. <br>
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        4. Incubate the fresh cultures at 37℃ until OD600 reach 0.2. <br>
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        A.Test the equality of the killing ability <br>
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        5.Construct the following 6 groups and then incubate the cultures at 37℃,shaking at 220rpm. For each group, 3 replicates are constructed, numbered from 1 to 3. Each replicate contains the bacteria with the same number. </div>
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    <div id="apDiv14"><img src="form1.jpg" alt="&lt;img src=&quot;form1.jpg&quot; width=&quot;863&quot; height=&quot;236&quot; /&gt;" width="631" height="236" />&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<img src="sodier.png" width="330" height="287"></div>
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    <div id="apDiv15">6.Take out 200ul cultures obtained from Procedure 5 and measure the fluorescent intensity and the amount of bacteria using flowcytometry after 15h.  <br>
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      <div id="apDiv16"><img src="form2.jpg" alt="&lt;img src=&quot;form2.jpg&quot; width=&quot;866&quot; height=&quot;301&quot; /&gt;" width="618" height="302" />&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;<img src="begger.png" width="344" height="202">&nbsp;</div>
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        B.Test if the killing ability is adequate <br>
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        5. Construct the following 8 groups and then incubate the cultures at 37℃, shaking at 220rpm. For each group, 3 replicates are constructed, numbered from 1 to 3. Each replicate contains the bacteria with the same number. All the medium contain 100ug/ml chloramphenicol. </div>
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    <div id="apDiv18">6.Take out 200ul cultures obtained from Procedure 5 and measure the fluorescent intensity and the amount of bacteria using flowcytometry after 15h. </div>
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    <div id="apDiv19">A.Test the equality of the killing ability:<br>
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1.warrior 1 kills itself but warrior doesn’t .We compare the amount of warriors in medium containing A+K with the amount of warriors in medium containing C. If the amount is apparently lower in the medium containing C, the warrior must have killed itself.We find that the amount of warrior 1 is apparently lower in C group, which means that warrior 1 kills itself.                                                    Figure1 shows the result. The ratio represents the amount of warriors in C medium divided by in A+K medium.(group4/group1,group 5/group 2) *A+K stands for ampicilin+kanamycin,C stands for chloramphenicol. </div>
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&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;       Fig.1 test of self-killing <br>
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    <div id="apDiv23"><img width="340" height="236" src="paiban_clip_image002_0001.gif" align="left" hspace="12" alt="killing ability is not equal"><br>
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              &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Fig.2 test the equality of killing</div>
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<div id="apDiv22">2.The killing ability of the two warriors are not equal We calculate the ratio of warrior 1/warrior 2 in group 3 and group 6. If the ratios are alike, then the killing ability of the warriors are similar.    Figure 2 shows the result.We can see that in the medium containing A+K, the amount of the warriors from two sides are roughly equal. At the same time, in the medium containing C, the amount of warrior 2 is much larger.This is easy to explain as not only does warrior 2 kills warrior 1, but warrior 1 kills itself as well.                                    *A+K stands for ampicilin+kanamycin,C stands for chloramphenicol. </div>
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<div id="apDiv21">IV.Results: </div>
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<div id="apDiv24">B.Test if the killing ability is adequate  <br>
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  &nbsp;&nbsp;&nbsp;We calculate the ratio of beggar/warrior in each group and the results can be seen from the figures.<br>
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&nbsp;&nbsp; We can see that when we mix warrior1 with beggar 2, the amount of beggar 2 after 15h is much more than the amount of warrior 1.This may partly because warrior1 kills itself and partly because its killing ability towards beggar 2 is weak.<br>
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&nbsp;&nbsp; In contrast with the result above, we find that when we mix beggar1 and warrior 2 together, the amount of warrior2 surely surpasses that of beggar1. Besides, the killing ability of warrior2 is adequate. The killing ability is neither too strong                                                                      when the ratio is 1:1, nor is it too weak when the ratio is 1:10.    <br>
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&nbsp;&nbsp; Furthermore, we compared the amount of beggars when they are cultured alone with the amount of beggars when they are cultured with warriors. We calculated the amount of beggars in group 7/group 8 to see the change in beggar1 and in group11/group12 to see the change in beggar2.From  figure 4, we can see that the amount of beggar1 drops drastically when it is cultured with warrior2,                                                                which means that warrior2 really work. As the amount of beggar2 does not have an apparent change, we may say that warrior1’s killing ability is surely too weak. </div>
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<div id="apDiv25"><img width="256" height="172" src="paiban_clip_image002_0003.gif" align="left" hspace="12" alt="w2_b1"><img width="246" height="167" src="paiban_clip_image004_0001.gif" align="left" hspace="12" alt="w1_b2"><br>
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<h4> What should we do for our demonstration?</h4>
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<h5> Standard teams </h5>
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If you have built a proof of concept system, you can demonstrate it working under real world conditions. If you have built a biological device that is intended to be a sensor, can you show it detecting whatever it is intended to sense. If it is intended to work in the field, you can show how this might work using a simulated version in the lab, or a simulation of your device in the field.<strong> Please note biological materials must not be taken out of the lab</strong>.
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<h5> Special track teams </h5>
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Special track teams can achieve this medal criterion by bringing their work to the Jamboree and showcasing it in the track event. Art & Design, Measurement, Hardware and Software tracks will all have showcase events at the Giant Jamboree.<strong> Please note biological materials must not be taken out of the lab</strong>.
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Revision as of 12:05, 21 October 2017

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