Difference between revisions of "Team:Ecuador/Parts"

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{{Ecuador}}
 
{{Ecuador}}
 
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<h1>Parts</h1>
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<p>Each team will make new parts during iGEM and will submit them to the Registry of Standard Biological Parts. The iGEM software provides an easy way to present the parts your team has created. The <code>&lt;groupparts&gt;</code> tag (see below) will generate a table with all of the parts that your team adds to your team sandbox.</p>
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  height: 300px;
−
<p>Remember that the goal of proper part documentation is to describe and define a part, so that it can be used without needing to refer to the primary literature. Registry users in future years should be able to read your documentation and be able to use the part successfully. Also, you should provide proper references to acknowledge previous authors and to provide for users who wish to know more.</p>
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<h3>Note</h3>
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<p>Note that parts must be documented on the <a href="http://parts.igem.org/Main_Page"> Registry</a>. This page serves to <i>showcase</i> the parts you have made. Future teams and other users and are much more likely to find parts by looking in the Registry than by looking at your team wiki.</p>
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</head>
  
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<div class="column two_thirds_size">
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<body>
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<div class="highlight decoration_B_full">
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<div class="ec--modeling--container">
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<div class="ec--first--section">
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<h3>Adding parts to the registry</h3>
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<div class="ec--main--title">
−
<p>You can add parts to the Registry at our <a href="http://parts.igem.org/Add_a_Part_to_the_Registry">Add a Part to the Registry</a> link.</p>
+
<a name="INTER"  style="text-decoration: none;"><div class="ec--h1">INTERLAB</div></a>
−
 
+
</div>
−
<p>We encourage teams to start completing documentation for their parts on the Registry as soon as you have it available. The sooner you put up your parts, the better you will remember all the details about your parts. Remember, you don't need to send us the DNA sample before you create an entry for a part on the Registry. (However, you <b>do</b> need to send us the DNA sample before the Jamboree. If you don't send us a DNA sample of a part, that part will not be eligible for awards and medal criteria.)</p>
+
<center>
−
<div class="button_link">
+
  <p><img width="385" height="244" src="https://static.igem.org/mediawiki/2018/d/d6/T--ECUADOR--Modeling.jpeg"></p></center>
−
<a href="http://parts.igem.org/Add_a_Part_to_the_Registry">
+
<div class="ec--modeling--items">
−
ADD PARTS
+
<ul>
−
</a>
+
<li><a href="#WWD">WHAT DID WE DO?</a></li>
−
</div>
+
<li><a href="#HWD">HOW DID WE DO IT?</a></li>
−
 
+
<li><a href="#BIO">BioBricks</a></li>
−
</div>
+
<li><a href="#RES">RESULTS</a></li>
−
</div>
+
<li><a href="#EXP">EXPERIENCE</a></li>
−
 
+
<li><a href="#ACK">ACKNOWLEDGEMENT</a></li>
−
 
+
</ul>
−
 
+
</div>
−
<div class="column third_size">
+
−
<div class="highlight decoration_A_full">
+
</div>
−
<h3>Inspiration</h3>
+
−
<p>We have a created  a <a href="http://parts.igem.org/Well_Documented_Parts">collection of well documented parts</a> that can help you get started.</p>
+
<div class="ec--nau--content">
−
 
+
  <a name="WWD"><div class="ec--h2">What did we do?</div></a>
−
<p> You can also take a look at how other teams have documented their parts in their wiki:</p>
+
<div class="ec--p">We tried to reduce the variability in the measurement of GFP expression for this reason we used a direct method to determine the mean expression level of GFP per cel</div>
−
<ul>
+
−
<li><a href="https://2014.igem.org/Team:MIT/Parts"> 2014 MIT </a></li>
+
  <a name="HWD"><div class="ec--h2">How did we do it?</div></a>
−
<li><a href="https://2014.igem.org/Team:Heidelberg/Parts"> 2014 Heidelberg</a></li>
+
<div class="ec--p">In order to avoid distortions in the results due to differences in the experimentation processes, we follow the procedures described in the iGEM protocol (<a href="https://static.igem.org/mediawiki/2018/0/09/2018_InterLab_Plate_Reader_Protocol.pdf" style="text-decoration: none">Plate reader protocol</a>, <a href="http://parts.igem.org/Help:Protocols/Competent_Cells" style="text-decoration: none">Competent cells</a>, <a href="http://parts.igem.org/Help:Protocols/Transformation" style="text-decoration: none">Transformation</a>)l</div>
−
<li><a href="https://2014.igem.org/Team:Tokyo_Tech/Parts">2014 Tokyo Tech</a></li>
+
−
</ul>
+
  <div class="ec--h3">Calibration</div>
−
</div>
+
<div class="ec--p">Before starting the experiments, we performed the OD600 calibration measurements following the LUDOX protocol, the particle Estandar Curve with the Microsphere Protocol, and the fluorescence standard curve with the Fluorescein Protocol. This allowed us to familiarize ourselves with the plate reader required for subsequent measurements. The plate reader that we used had the following characteristics:<br />
−
</div>
+
  Label: BioTek / Model: Cytation 5 / Emission: 528 / Excitation: 485 / Optic position: Top / Temperature: 25 ºC / Shake: 425 cpm for 30 sec before every measurement.
−
 
+
</div>
−
 
+
−
<div class="clear extra_space"></div>
+
<div class="ec--h3">Cell measurement</div>
−
 
+
<div class="ec--p">
−
 
+
We started by transforming E. coli DH5 alpha with the plasmid described in the protocol, and after incubation and growth in LB-medium, we diluted the culture to Abs600 0.2 and proceeded  with the samples.
−
 
+
</div>
−
 
+
−
<div class="column full_size">
+
<div class="ec--h3">Colony units per 0.1 OD600 E. coli cultures</div>
−
 
+
<div class="ec--p">
−
<h3>What information do I need to start putting my parts on the Registry?</h3>
+
In order to determine the cell concentration of the culture we followed the CFU protocol that basically consisted in diluting a culture to certain absorbance, measure the OD600 that has to be in a value of 0.1, and make serial dilutions to later count the colonies and calculate the CFU/mL.
−
<p>The information needed to initially create a part on the Registry is:</p>
+
</div>
−
<ul>
+
−
<li>Part Name</li>
+
  <a name="BIO"><div class="ec--h2">BioBricks</div></a>
−
<li>Part type</li>
+
−
<li>Creator</li>
+
<div class="ec--biob--cont">
−
<li>Sequence</li>
+
<div id="bigBox">
−
<li>Short Description (60 characters on what the DNA does)</li>
+
−
<li>Long Description (Longer description of what the DNA does)</li>
+
<div id="leftBox">
−
<li>Design considerations</li>
+
    <table border="0" cellspacing="8px" cellpadding="0">
−
</ul>
+
          <tr>
−
 
+
            <td width="148" valign="top"><center ><strong>Device</strong><strong> </strong></center></td>
−
<p>
+
            <td width="132" valign="top"><center><strong>Part Number</strong><strong> </strong></center></td>
−
We encourage you to put up <em>much more</em> information as you gather it over the summer. If you have images, plots, characterization data and other information, please also put it up on the part page. </p>
+
              </tr>
−
 
+
          <tr>
−
</div>
+
            <td width="148" valign="top"><p id="label-neg-c" class="ec--inter-item">Negative control</p></td>
−
 
+
            <td width="132" valign="top"><p>BBa_R0040 </p></td>
−
 
+
              </tr>
−
<div class="clear extra_space"></div>
+
          <tr>
−
<div class="line_divider"></div>
+
            <td width="148" valign="top"><p id="type"class="ec--inter-item">Positive control </p></td>
−
<div class="clear extra_space"></div>
+
            <td width="132" valign="top"><p>BBa_I20270 </p></td>
−
 
+
              </tr>
−
<div class="column full_size">
+
          <tr>
−
<h3>Part Table </h3>
+
            <td width="148" valign="top"><p id="label-td1" class="ec--inter-item">Test Device 1 </p></td>
−
 
+
            <td width="132" valign="top"><p>BBa_J364000 </p></td>
−
<p>Please include a table of all the parts your team has made during your project on this page. Remember part characterization and measurement data must go on your team part pages on the Registry. </p>
+
              </tr>
−
 
+
          <tr>
−
</html>
+
            <td width="148" valign="top"><p id="label-td2" class="ec--inter-item">Test Device 2 </p></td>
−
<groupparts>iGEM18 Ecuador</groupparts>
+
            <td width="132" valign="top"><p>BBa_J364001 </p></td>
−
<html>
+
              </tr>
−
</div>
+
          <tr>
−
 
+
            <td width="148" valign="top"><p id="label-td3" class="ec--inter-item">Test Device 3 </p></td>
−
 
+
            <td width="132" valign="top"><p>BBa_J364002 </p></td>
−
 
+
              </tr>
−
 
+
          <tr>
−
</html>
+
            <td width="148" valign="top"><p id="label-td4" class="ec--inter-item">Test Device 4 </p></td>
 +
            <td width="132" valign="top"><p>BBa_J364007 </p></td>
 +
              </tr>
 +
          <tr>
 +
            <td width="148" valign="top"><p id="label-td5" class="ec--inter-item">Test Device 5 </p></td>
 +
            <td width="132" valign="top"><p>BBa_J364008 </p></td>
 +
              </tr>
 +
          <tr>
 +
            <td width="148" valign="top"><p id="label-td6" class="ec--inter-item">Test Device 6 </p></td>
 +
            <td width="132" valign="top"><p>BBa_J364009 </p></td>
 +
              </tr>
 +
            </table>
 +
  </div>
 +
<div id="rightBox">
 +
<div class="ec--biobr--location--container">
 +
<table width="338" border="0" cellpadding="0" cellspacing="8px">
 +
  <tr>
 +
<td width="330" valign="top"><p align="center"><strong>Structure<br>
 +
(Point with the cursor on the device)
 +
</strong></p></td>
 +
  </tr>
 +
</table>
 +
<div class="ec--biobr--separator"></div>
 +
<div class="loc-neg-c">
 +
 +
<p><img width="359" height="75" src="https://static.igem.org/mediawiki/2018/c/c9/T--ECUADOR--L001.png"> </p>
 +
<br>
 +
<p>length: 54 bp </p>
 +
  </div>
 +
<div class="textdis" id="pos_c">
 +
 +
<p><img width="195" height="70" src="https://static.igem.org/mediawiki/2018/b/b5/T--ECUADOR--L002.png"></p>
 +
<br>
 +
<p>length: 919 bp</p>
 +
</div>
 +
<div class="loc-td1">
 +
 +
<p><img width="195" height="51" src="https://static.igem.org/mediawiki/2018/8/86/T--ECUADOR--L003.png" > </p>
 +
<br>
 +
<p>length: 918 bp </p>
 +
</div>
 +
<div class="loc-td2">
 +
 +
<p><img width="189" height="54" src="https://static.igem.org/mediawiki/2018/c/c8/T--ECUADOR--L004.png" ></p>
 +
<br>
 +
<p>length: 918 bp</p>
 +
</div>
 +
<div class="loc-td3">
 +
 +
<p><img width="197" height="63" src="https://static.igem.org/mediawiki/2018/4/43/T--ECUADOR--L005.png" > </p>
 +
<br>
 +
<p>length: 918 bp </p>
 +
</div>
 +
<div class="loc-td4">
 +
 +
<p><img width="196" height="52" src="https://static.igem.org/mediawiki/2018/5/53/T--ECUADOR--L006.png"></p>
 +
<br>
 +
<p>length: 918 bp</p>
 +
</div>
 +
<div class="loc-td5">
 +
 +
<p><img width="188" height="64" src="https://static.igem.org/mediawiki/2018/9/97/T--ECUADOR--L007.png" > </p>
 +
<br>
 +
<p>length: 918 bp </p>
 +
</div>
 +
<div class="loc-td6">
 +
 +
<p><img width="194" height="55" src="https://static.igem.org/mediawiki/2018/0/02/T--ECUADOR--L008.png"></p>
 +
<br>
 +
<p>length: 918 bp</p>
 +
</div>
 +
    </div>
 +
</div>
 +
  </div>
 +
  </div>
 +
 +
<a name="RES"><div class="ec--h2">Results</div></a>
 +
  <div class="ec--h3">Particle standard curves</div>
 +
<div class="ec--img--wwd--cont">
 +
<img src="https://static.igem.org/mediawiki/2018/3/30/T--ECUADOR--A-PSC.1.png" width="537" height="307" alt=""/>
 +
</div>
 +
<div class="ec--img--wwd--cont">
 +
<img src="https://static.igem.org/mediawiki/2018/1/1d/T--ECUADOR--A-PSC.2.png" width="534" height="303" alt=""/>
 +
</div>
 +
 +
<div class="ec--h3">Fluorescein standard curves</div>
 +
<div class="ec--img--wwd--cont">
 +
    <p><img src="https://static.igem.org/mediawiki/2018/7/7c/T--ECUADOR--A-FSC.1.png" width="534" height="303" alt=""/><br></p>
 +
  </div>
 +
              <div class="ec--img--wwd--cont">
 +
  <img src="https://static.igem.org/mediawiki/2018/c/cf/T--ECUADOR--A-FSC.2.png" width="534" height="303" alt=""/><br>
 +
  </div>
 +
                <div class="ec--h3">Converting between absorbance of cells to absorbance of a known concentration of beads</div>
 +
<div class="ec--img--wwd--cont">
 +
              <p><img src="https://static.igem.org/mediawiki/2018/6/64/T--ECUADOR--A-NA600.png" width="534" height="303" alt=""/></p>
 +
</div>
 +
  <div class="ec--h3">Counting colony-forming units (CFUs) from the sample</div>
 +
<div class="ec--img--wwd--cont">
 +
              <p><img src="https://static.igem.org/mediawiki/2018/2/2e/T--ECUADOR--A-RBT.png" width="534" height="303" alt=""/></p>
 +
</div>
 +
 +
  <a name="EXP"><div class="ec--h2">EXPERIENCE</div></a>
 +
</a>
 +
              <div class="ec--img--wwd--cont">
 +
                <center><img src="https://svgsilh.com/svg/2324013-2196f3.svg" width="309" height="175" alt=""/></center>
 +
              </div>
 +
              <div class="ec--p">The interlab protocols were easy to understand and simple. We recommend the following:</div>
 +
              <ul>
 +
                <li>
 +
                  <div class="ec--p">Before each measurement, shake the plates in the equipment since the particles tend to precipitate in the well.</div>
 +
                </li>
 +
                <li dir="ltr">
 +
                  <div class="ec--p">If the iGEM equipment does not have the plate reader in the same place as your laboratory, they must take care of their samples in continuous refrigeration during transport and carry out the activities with time.</div>
 +
                </li>
 +
              </ul>
 +
              <div class="ec--p">We learned many things in the interlab challenge: the protocols, the management of the plate reader and how it can be used in later processes of our project. It also helped us strengthen cooperation with other universities.</div>
 +
 +
              <a name="ACK" style="text-decoration: none;">
 +
              <div class="ec--h2">ACKNOWLEDGEMENT</div>
 +
              </a>
 +
<div class="ec--p">
 +
As a team, we would like to thank the Biomedicine team of Universidad Tecnica Equinoccial and especially Linda  Guaman, member of their research team, for allowing us to use their measurement  equipment, which were essential for the completion of the Interlab.</div>
 +
 +
</div>
 +
</div>
 +
 +
</body>
 +
 +
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Revision as of 09:04, 17 October 2018

iGEM ECUADOR
C-lastin, Interlab
<div class="ec--h1">INTERLAB</div>

<div class="ec--h2">What did we do?</div>
We tried to reduce the variability in the measurement of GFP expression for this reason we used a direct method to determine the mean expression level of GFP per cel
<div class="ec--h2">How did we do it?</div>
In order to avoid distortions in the results due to differences in the experimentation processes, we follow the procedures described in the iGEM protocol (Plate reader protocol, Competent cells, Transformation)l
Calibration
Before starting the experiments, we performed the OD600 calibration measurements following the LUDOX protocol, the particle Estandar Curve with the Microsphere Protocol, and the fluorescence standard curve with the Fluorescein Protocol. This allowed us to familiarize ourselves with the plate reader required for subsequent measurements. The plate reader that we used had the following characteristics:
Label: BioTek / Model: Cytation 5 / Emission: 528 / Excitation: 485 / Optic position: Top / Temperature: 25 ºC / Shake: 425 cpm for 30 sec before every measurement.
Cell measurement
We started by transforming E. coli DH5 alpha with the plasmid described in the protocol, and after incubation and growth in LB-medium, we diluted the culture to Abs600 0.2 and proceeded with the samples.
Colony units per 0.1 OD600 E. coli cultures
In order to determine the cell concentration of the culture we followed the CFU protocol that basically consisted in diluting a culture to certain absorbance, measure the OD600 that has to be in a value of 0.1, and make serial dilutions to later count the colonies and calculate the CFU/mL.
<div class="ec--h2">BioBricks</div>
Device
Part Number

Negative control

BBa_R0040

Positive control

BBa_I20270

Test Device 1

BBa_J364000

Test Device 2

BBa_J364001

Test Device 3

BBa_J364002

Test Device 4

BBa_J364007

Test Device 5

BBa_J364008

Test Device 6

BBa_J364009

Structure
(Point with the cursor on the device)


length: 54 bp


length: 919 bp


length: 918 bp


length: 918 bp


length: 918 bp


length: 918 bp


length: 918 bp


length: 918 bp

<div class="ec--h2">Results</div>
Particle standard curves
Fluorescein standard curves



Converting between absorbance of cells to absorbance of a known concentration of beads

Counting colony-forming units (CFUs) from the sample

<div class="ec--h2">EXPERIENCE</div>
The interlab protocols were easy to understand and simple. We recommend the following:
  • Before each measurement, shake the plates in the equipment since the particles tend to precipitate in the well.
  • If the iGEM equipment does not have the plate reader in the same place as your laboratory, they must take care of their samples in continuous refrigeration during transport and carry out the activities with time.
We learned many things in the interlab challenge: the protocols, the management of the plate reader and how it can be used in later processes of our project. It also helped us strengthen cooperation with other universities.
<div class="ec--h2">ACKNOWLEDGEMENT</div>
As a team, we would like to thank the Biomedicine team of Universidad Tecnica Equinoccial and especially Linda Guaman, member of their research team, for allowing us to use their measurement equipment, which were essential for the completion of the Interlab.
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