Difference between revisions of "Team:JNFLS/Parts"

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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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<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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<h3>Our Parts </h3>
  
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<p>Our project is to develop a biosensor based on the aptamer target on the HCV core protein, using competition based rolling amplification. This device could be used for the detection of trace HCV virus in unpaid donation blood test and blood transfusion in the clinic, which could shorten the window period of HCV detection. In order to achieve enough HCV C protein expressed in Prokaryotes, we constructed some recombinant plasmids of HCV C gene, some of them are truncated or/and optimized, some of them are tagged with TEE and 6×His marker. We cloned all these modified HCV C genes into pSB1C3 plasmid backbone to produce new BioBrick parts, which are compatible with RFC 10. All those parts worked well, particularly BBa_K2608004, which was selected to use in our project. </p>
  
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<p>HCV is a single strand RNA virus with a coat. Core antigen is the main protein of nucleocapsid. It is an important part of the virus particle. Core protein can control the virus itself, as well as the interactions of virus and host cells. After signal peptide was inserted to the 5’-UTR of HCV C gene, the expressed fusion protein can be secreted to the outside of the cell, with the ability of self-assembly into the virus particle. Ii is a very good tool for HCV research. All the part genes were inserted into the vector plasmid pColdⅡ, showed as below.</p>
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<img src="https://static.igem.org/mediawiki/2018/thumb/a/a0/T--JNFLS--baobao.jpg/784px-T--JNFLS--baobao.jpg.png"styel="width:0.1%">
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<b>Fig.1 Map of pCold Ⅱvector shows TEE and His tag are inserted to the upstream of multiple cloning sites.</b>
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<img src="https://static.igem.org/mediawiki/2018/f/fb/T--JNFLS--chongz.jpg"styel="width:10%">
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<b>Fig.2 Diagram of pCold Ⅱvector shows the cloning site of HCV C gene insertion.</b>
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<p>We constructed 10 HCV core protein parts with different length and different tag, even different sequences due to the codon optimization. All the part information are summarized in the table below.</p>
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<img src="https://static.igem.org/mediawiki/2018/6/68/T--JNFLS--tb.png"styel="width:70%">
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<p>In these parts, the BBa_K2608004 was the shortest one which contains 360bp, encoding 120aa without tagging other sequence. It was also optimized and expressed very well, which showed as following.</p>
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<img src="https://static.igem.org/mediawiki/2018/b/b4/T--JNFLS--gf1.png"styel="width:70%">
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<b>Fig.3 The centrifuge pellets of bacteria shows the induced expression of fusion HCV C protein in E.coli</b>
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<img src="https://static.igem.org/mediawiki/2018/9/99/T--JNFLS--sds1.png"styel="width:70%">
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<b>Fig.4 SDS-PAGE result showing the expression of HCV C-O173 and HCV C-O120, which were marked by GFP</b>
  
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<h3>Adding parts to the registry</h3>
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<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>
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<p>You can view our parts here:</p>
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<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>
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<a href="http://parts.igem.org/Add_a_Part_to_the_Registry">
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ADD PARTS
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<h3>Inspiration</h3>
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<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>
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<p> You can also take a look at how other teams have documented their parts in their wiki:</p>
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<li><a href="https://2014.igem.org/Team:MIT/Parts"> 2014 MIT </a></li>
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<li><a href="https://2014.igem.org/Team:Heidelberg/Parts"> 2014 Heidelberg</a></li>
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<li><a href="https://2014.igem.org/Team:Tokyo_Tech/Parts">2014 Tokyo Tech</a></li>
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<h3>What information do I need to start putting my parts on the Registry?</h3>
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<p>The information needed to initially create a part on the Registry is:</p>
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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>
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<h3>Part Table </h3>
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<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>
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<groupparts>iGEM18 JNFLS</groupparts>
 
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Latest revision as of 15:39, 17 October 2018

Our Parts

Our project is to develop a biosensor based on the aptamer target on the HCV core protein, using competition based rolling amplification. This device could be used for the detection of trace HCV virus in unpaid donation blood test and blood transfusion in the clinic, which could shorten the window period of HCV detection. In order to achieve enough HCV C protein expressed in Prokaryotes, we constructed some recombinant plasmids of HCV C gene, some of them are truncated or/and optimized, some of them are tagged with TEE and 6×His marker. We cloned all these modified HCV C genes into pSB1C3 plasmid backbone to produce new BioBrick parts, which are compatible with RFC 10. All those parts worked well, particularly BBa_K2608004, which was selected to use in our project.

HCV is a single strand RNA virus with a coat. Core antigen is the main protein of nucleocapsid. It is an important part of the virus particle. Core protein can control the virus itself, as well as the interactions of virus and host cells. After signal peptide was inserted to the 5’-UTR of HCV C gene, the expressed fusion protein can be secreted to the outside of the cell, with the ability of self-assembly into the virus particle. Ii is a very good tool for HCV research. All the part genes were inserted into the vector plasmid pColdⅡ, showed as below.

Fig.1 Map of pCold Ⅱvector shows TEE and His tag are inserted to the upstream of multiple cloning sites.

Fig.2 Diagram of pCold Ⅱvector shows the cloning site of HCV C gene insertion.

We constructed 10 HCV core protein parts with different length and different tag, even different sequences due to the codon optimization. All the part information are summarized in the table below.

In these parts, the BBa_K2608004 was the shortest one which contains 360bp, encoding 120aa without tagging other sequence. It was also optimized and expressed very well, which showed as following.

Fig.3 The centrifuge pellets of bacteria shows the induced expression of fusion HCV C protein in E.coli

Fig.4 SDS-PAGE result showing the expression of HCV C-O173 and HCV C-O120, which were marked by GFP

You can view our parts here:

<groupparts>iGEM18 JNFLS</groupparts>