Difference between revisions of "Team:Jilin China/Composite Part"

Line 51: Line 51:
 
   <p>COMPOSITE PART</p>
 
   <p>COMPOSITE PART</p>
 
   <br />
 
   <br />
−
   <!--table>
+
   <table>
 
   <tr>
 
   <tr>
−
     <td><a href="#paragraph_1" class="clickwave">Absract</a></td>
+
     <td><a href="#paragraph_1" class="clickwave">Abstract</a></td>
−
     <td><a href="#paragraph_2" class="clickwave">Parts</a></td>
+
     <td><a href="#paragraph_2" class="clickwave">Thermosensor</a></td>
−
     <td><a href="#paragraph_3" class="clickwave">References</a></td>
+
     <td><a href="#paragraph_3" class="clickwave">sfGFP</a></td>
  
  
 
   </tr>
 
   </tr>
−
   </table-->
+
   </table>
 
   </div>
 
   </div>
 
    
 
    
Line 68: Line 68:
 
   <ul class="sidenav">
 
   <ul class="sidenav">
 
   <li><a href="#paragraph_1">Abstract</a></li>
 
   <li><a href="#paragraph_1">Abstract</a></li>
−
   <li><a href="#paragraph_2">Parts</a></li>
+
   <li><a href="#paragraph_2">Thermosensor</a></li>
−
   <li><a href="#paragraph_3">References</a></li>
+
   <li><a href="#paragraph_3">sfGFP</a></li>
 
   </ul>
 
   </ul>
 
   <section class="s2">
 
   <section class="s2">
 
   <ul>
 
   <ul>
−
    <li class="toBeDeletedAlert full-width" id="">
 
−
    <div>
 
−
      <h2>Editing Alert!</h2>
 
−
 
−
      <p>This page is used by the judges to evaluate your team for the <a href="https://2018.igem.org/Judging/Medals">medal criterion</a> or <a href="https://2018.igem.org/Judging/Awards"> award listed below</a>. </p>
 
−
 
−
      <h3>Note</h3>
 
−
      <p>This page should list all the composite parts your team has made during your project. You must add all characterization information for your parts on the Registry. You should not put characterization information on this page. Remember judges will only look at the first part in the list for the Best Basic Part award, so put your best part first!</p>
 
−
 
−
      <h3>Best Basic Part Special Prize</h3>
 
−
      <p> To be eligible for this award, this part must adhere to <a href="http://parts.igem.org/DNA_Submission">Registry sample submission guidelines</a> and have been sent to the Registry of Standard Biological Parts. If you have a part you wish to nominate your team for this <a href="https://2018.igem.org/Judging/Awards">special prize</a>, make sure you add your part number to your <a href="https://2018.igem.org/Judging/Judging_Form">judging form</a> and delete the box at the top of this page.
 
−
<br>
 
−
<b>Please note:</b> Judges will only look at the first part number you list, so please only enter ONE (1) part number in the judging form for this prize. </p>
 
−
    </div>
 
−
    </li>
 
 
     <li class="paragraph_1 start" id="paragraph_1">
 
     <li class="paragraph_1 start" id="paragraph_1">
 
     <div>
 
     <div>
−
       <h2>Overview</h2>
+
       <h2>Abstract</h2>
−
<p>This year, we added a total of xx composite parts, including cold-induced RNA thermosensors consisting of Anderson promoter J23104 and cspA mRNA, and a color-selection device designed for Goldengate assembly. We decided to use our composite part B123456 as a strong contender for the Best composite part. </p>
+
<p>This year, we added a total of 7 composite parts, including heat-inducible RNA-based thermosensor measurement device, heat-repressible RNA-based thermosensor measurement device, cold-inducible RNA-based thermosensor measurement device, cold-repressible RNA-based thermosensors measurement device, sfGFP_optimism measurement device, superfolder GFP measurement device and sfGFP(BbsI free)measurement device.</p>
 
     </div>
 
     </div>
 
     </li>
 
     </li>
 
     <li class="paragraph_2" id="paragraph_2">
 
     <li class="paragraph_2" id="paragraph_2">
 
     <div>
 
     <div>
−
      <h2>Color-selection Device for GoldenGate Assembly</h2>
+
<h2>Thermosensor measurement device</h2>
−
<p>We use the color-selection device for goldengate assembly to select the right  plasmids based on the color change of monoclonal colonies. The Goldengate assembly is a process of seamless assembly by using IIs-type restriction endonucleases. In our project, we decided to use goldengate assembly to construct our vast number of measurement devices. However, for the successful construction of plasmids, the selection has become a key step in limiting the speed of construction. In order to solve this problem, Jilin_China designed the following construction device. </p>
+
<table width="200" border="1">
−
<p>Our construction device includes a promoter J23101 and chromoprotein tspurple, which can be replaced by the BbsI restriction endonuclease, cjblue that can be replaced by the BsaI restriction endonuclease, conserved sequence and reporter protein sfGFP. Such a composition can be realized that when both the replacement part I and the replacement part II are not replaced, the colony exhibits the color of the tspurple, and when the replacement part I is replaced with another promoter by using BbsI, the colony appears blue, and the unsuccessful colony appears Purple. When BsaI was used to replace Part II, the colony showed the original milky white color of the bacteria, and the blue color colony is unsuccess. The specific color changes are as follows: </p>
+
  <tbody>
−
<div align="center">
+
    <tr>
−
<img src="https://static.igem.org/mediawiki/2018/2/25/T--Jilin_China--Parts--Composite_Part--color_table.png" align="center"/>
+
      <td>Part Name</td>
 +
      <td>Part Number</td>
 +
    </tr>
 +
  <tr>
 +
      <td>Heat-inducible RNA-based thermosensor measurement device</td>
 +
      <td>BBa_K2541405</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Heat-repressible RNA-based thermosensor measurement device</td>
 +
      <td>BBa_K2541406</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Cold-repressible RNA-based thermosensor measurement device</td>
 +
      <td>BBa_K2541407</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Cold-inducible RNA-based thermosensor measurement device</td>
 +
      <td>BBa_K2541408</td>
 +
    </tr>
 +
  </tbody>
 +
</table>
 +
<p>The thermosensor sequence is constructed on the pSB1C3 vector by Golden Gate assembly. The measurement device is composed of Anderson promotor (BBa_J23104), thermosensor (BBa_K2541051), sfGFP_optimism (BBa_K2541400) and double terminator (BBa_B0010 and BBa_B0012). We select a constitutive Anderson promoter J23104 as an appropriate promoter by pre-experiment. The sfGFP_optimism has faster folding speed and higher fluorescence intensity. The double terminator can reduce leakage (Figure 1). We characterized RNA-based thermosensors in E.coli DH5a.</p>
 +
<img src="http://parts.igem.org/File:Measurement_device_figure3.png" />
 +
<center>Figure 3. Diagram of the measurement device.</center>
 +
 
</div>
 
</div>
−
<p>Through the visible color changes, we can select successful colonies from the culture dish. In this way, we can eliminate the gel midi purification work, reduce the workload and increase the work efficiency compared with the traditional construction method. There is also a direction in the choice of picking up monoclonal colonies. </p>
 
−
<p>After actual experiments, we can successfully build more than 200 plasmids in one day, and the sequencing success rate is over 95%, which is a very exciting result. </p>
 
−
<p>Here, we provide a method for efficient and seamless assembly. Users can add more replacements parts using different Type II restriction endonucleases according to their actual conditions, or add a conserved sequence between the replacement parts. </p>
 
−
<p>Our strength lies in combining high-efficiency goldengate assembly with chromoprotein selection for greater accuracy and productivity. </p>
 
−
<p>If you want to know more about our construction, please visit our construction page, where we will detail the principles of Goldengate. </p>
 
−
<center><a href="https://2018.igem.org/Team:Jilin_China/Improve">>Click here to explore more information! <</a></center>
 
−
<p>We have recorded the detailed protocol of the goldengate assembly on the protocol page. Other users can refer to our protocol to design their own experiments (the amount of each added reagent in the protocol has been reduced to a minimum, it is not recommended to continue to reduce the amount. At the same time, when actually selecting other types of Type IIs restriction endonucleases, you should test whether the enzyme can work normally in T4 buffer. We have tested T4 ligase and T4 buffer from NEB and Takara to use our system, others has not tested yet.) .</p>
 
−
    </div>
 
 
     </li>
 
     </li>
 
     <li class="paragraph_3" id="paragraph_3">
 
     <li class="paragraph_3" id="paragraph_3">
 
     <div>
 
     <div>
−
       <h2>Cold-induced RNA Thermosensor Parts</h2>
+
       <h2>sfGFP Measurement Device</h2>
 
<p>In addition, we combined our cold-induced RNA thermosensors (cspA mRNA) with the promoter J23104 as a composite part for uploading, which is more conducive to other users directly, users can also only use a cold-induced RNA thermosensor according to their own needs.</p>
 
<p>In addition, we combined our cold-induced RNA thermosensors (cspA mRNA) with the promoter J23104 as a composite part for uploading, which is more conducive to other users directly, users can also only use a cold-induced RNA thermosensor according to their own needs.</p>
 +
<table width="200" border="1">
 +
  <tbody>
 +
    <tr>
 +
      <td>Part Name</td>
 +
      <td>Part Number</td>
 +
    </tr>
 +
    <tr>
 +
      <td>sfGFP_optimism measurement device</td>
 +
      <td>BBa_K2541402</td>
 +
    </tr>
 +
    <tr>
 +
      <td>superfolder GFP measurement device</td>
 +
      <td>BBa_K2541403</td>
 +
    </tr>
 +
    <tr>
 +
      <td>sfGFP(BbsI free) measurement device</td>
 +
      <td>BBa_K2541404</td>
 +
    </tr>
 +
  </tbody>
 +
</table>
 +
<h3>Introduction</h3>
 +
<p>They are three measurement device for sfGFP_optimism (BBa_K2541400), superfolder GFP (BBa_I746916) and sfGFP(BbsI free)(BBa_K2541401) fluorescence intensity, which is composed of Anderson promoter (BBa_J23104), RBS (BBa_B0034), sfGFP and double terminator (BBa_B0010 and BBa_B0012).</p>
 
     </div>
 
     </div>
 
     </li>
 
     </li>

Revision as of 18:16, 15 October 2018

COMPOSITE PART


Abstract Thermosensor sfGFP

Composite Part

  • Abstract

    This year, we added a total of 7 composite parts, including heat-inducible RNA-based thermosensor measurement device, heat-repressible RNA-based thermosensor measurement device, cold-inducible RNA-based thermosensor measurement device, cold-repressible RNA-based thermosensors measurement device, sfGFP_optimism measurement device, superfolder GFP measurement device and sfGFP(BbsI free)measurement device.

  • Thermosensor measurement device

    Part Name Part Number
    Heat-inducible RNA-based thermosensor measurement device BBa_K2541405
    Heat-repressible RNA-based thermosensor measurement device BBa_K2541406
    Cold-repressible RNA-based thermosensor measurement device BBa_K2541407
    Cold-inducible RNA-based thermosensor measurement device BBa_K2541408

    The thermosensor sequence is constructed on the pSB1C3 vector by Golden Gate assembly. The measurement device is composed of Anderson promotor (BBa_J23104), thermosensor (BBa_K2541051), sfGFP_optimism (BBa_K2541400) and double terminator (BBa_B0010 and BBa_B0012). We select a constitutive Anderson promoter J23104 as an appropriate promoter by pre-experiment. The sfGFP_optimism has faster folding speed and higher fluorescence intensity. The double terminator can reduce leakage (Figure 1). We characterized RNA-based thermosensors in E.coli DH5a.

    Figure 3. Diagram of the measurement device.

  • sfGFP Measurement Device

    In addition, we combined our cold-induced RNA thermosensors (cspA mRNA) with the promoter J23104 as a composite part for uploading, which is more conducive to other users directly, users can also only use a cold-induced RNA thermosensor according to their own needs.

    Part Name Part Number
    sfGFP_optimism measurement device BBa_K2541402
    superfolder GFP measurement device BBa_K2541403
    sfGFP(BbsI free) measurement device BBa_K2541404

    Introduction

    They are three measurement device for sfGFP_optimism (BBa_K2541400), superfolder GFP (BBa_I746916) and sfGFP(BbsI free)(BBa_K2541401) fluorescence intensity, which is composed of Anderson promoter (BBa_J23104), RBS (BBa_B0034), sfGFP and double terminator (BBa_B0010 and BBa_B0012).