Difference between revisions of "Part:BBa K3187012"
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<partinfo>BBa_K3187012 short</partinfo>
 
<partinfo>BBa_K3187012 short</partinfo>
  
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<h2> pTeTW3con2-ptet-sfGFP -StrepTag-SP—CP+LPETGG-pT7:</h2>
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<       <li> Name: pTeTW3con2-ptet-sfGFP-StrepTag-SP—CP+LPETGG-pT7</li>
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<p>
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    <ul>
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        <li> Name: pTeTW3con2-ptet-sfGFP-StrepTag-SP—CP+LPETGG-pT7</li>
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         <li> Size [bp]: 2932 </li>
 
         <li> Size [bp]: 2932 </li>
 
         <li> Size [kDa]: 46.1 (sfGFP – Strep-Tag – Scaffold Protein) + 48.9 (Coat Protein - Strep-Tag - LPETGG) </li>
 
         <li> Size [kDa]: 46.1 (sfGFP – Strep-Tag – Scaffold Protein) + 48.9 (Coat Protein - Strep-Tag - LPETGG) </li>
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<p>
 
<p>
 
     This composite part is a dual expression plasmid with pTeTW3con2 as backbone and two convergent expression sites. The tetA promoter is
 
     This composite part is a dual expression plasmid with pTeTW3con2 as backbone and two convergent expression sites. The tetA promoter is
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     inducible with anhydrotetracycline (AHT) and the site <a href="https://parts.igem.org/Part:BBa_K3187043
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     inducible with anhydrotetracycline (AHT) and the site <a href="https://parts.igem.org/Part:BBa_K3187043" target="_blank">(BBa_K3187043)</a>encodes the fusion protein of sfGFP, Strep-Tag and scaffold
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    " target="_blank">(BBa_K3187043)</a>encodes the fusion protein of sfGFP, Strep-Tag and scaffold
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     protein. The T7 promoter can be induced by isopropyl β-D-1-thiogalactopyranoside (IPTG) and the site<a href="https://parts.igem.org/Part:BBa_K3187044" target="_blank">(BBa_K3187044
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     protein. The T7 promoter can be induced by isopropyl β-D-1-thiogalactopyranoside (IPTG) and the site<a href="https://parts.igem.org/Part:BBa_K3187044
+
−
    " target="_blank">(BBa_K3187044
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         )</a> encodes the
 
         )</a> encodes the
 
     fusion protein of coat protein, Strep-Tag and LPETGG. The composite part can therefore be used for the production of
 
     fusion protein of coat protein, Strep-Tag and LPETGG. The composite part can therefore be used for the production of
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of sfGFP with the scaffold protein the VLPs will contain sfGFP as a cargo on the inside.
 
of sfGFP with the scaffold protein the VLPs will contain sfGFP as a cargo on the inside.
 
<p>
 
<p>
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     The coat fusion protein also includes a Strep-Tag <a href="https://parts.igem.org/Part:BBa_K3187025
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     The coat fusion protein also includes a Strep-Tag <a href="https://parts.igem.org/Part:BBa_K3187025" target="_blank">(BBa_K3187025
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  " target="_blank">(BBa_K3187025
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         )</a> for purification and a LPETGG tag<a href="https://parts.igem.org/Part:BBa_K3187019" target="_blank">(BBa_K3187019
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         )</a> for purification and a LPETGG tag<a href="https://parts.igem.org/Part:BBa_K3187019
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−
    " target="_blank">(BBa_K3187019
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         )</a> . As a result of the fusion of
 
         )</a> . As a result of the fusion of
 
     the LEPTGG tag with the coat protein the VLPs will present the tag on the outside. This tag is the recognition
 
     the LEPTGG tag with the coat protein the VLPs will present the tag on the outside. This tag is the recognition
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     sequence of Sortase A7M<a href="https://parts.igem.org/Part:BBa_K3187028
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     sequence of Sortase A7M<a href="https://parts.igem.org/Part:BBa_K3187028" target="_blank">(BBa_K3187028
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    " target="_blank">(BBa_K3187028
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         )</a> which catalyzes a covalent bond of the LPETGG and a poly G tag. On this basis the VLPs can
 
         )</a> which catalyzes a covalent bond of the LPETGG and a poly G tag. On this basis the VLPs can
 
     be
 
     be
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<p>
 
<p>
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     The expression levels of both sites were characterized using the <a href="https://parts.igem.org/Part:BBa_K3187028
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     The expression levels of both sites were characterized using the <a href="https://parts.igem.org/Part:BBa_K3187028" target="_blank">BBa_K3187011</a>.  
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    " target="_blank">BBa_K3187011</a>.  
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</p>
 
</p>
 +
  
  

Revision as of 12:03, 17 October 2019


pTet sfGFP-P22 Bacteriophage Scaffolding Protein x Coat Protein pT7 (Convergent)

<
  • Name: pTeTW3con2-ptet-sfGFP-StrepTag-SP—CP+LPETGG-pT7
  • Size [bp]: 2932
  • Size [kDa]: 46.1 (sfGFP – Strep-Tag – Scaffold Protein) + 48.9 (Coat Protein - Strep-Tag - LPETGG)
  • Origin: Synthetic?
  • Components: tetA promoter, T7 promoter, T7 terminator, Lac Operator, RBS, Coat Protein, Scaffold Protein, sfGFP, LPETGG, Strep-Tag II
  • Feature: Production of P22 Virus-like particles with a modifiable surface and sfGFP as cargo 
    •    </ul>

    </p>


    Introduction:

    This composite part is a dual expression plasmid with pTeTW3con2 as backbone and two convergent expression sites. The tetA promoter is inducible with anhydrotetracycline (AHT) and the site <a href="https://parts.igem.org/Part:BBa_K3187043" target="_blank">(BBa_K3187043)</a>encodes the fusion protein of sfGFP, Strep-Tag and scaffold protein. The T7 promoter can be induced by isopropyl β-D-1-thiogalactopyranoside (IPTG) and the site<a href="https://parts.igem.org/Part:BBa_K3187044" target="_blank">(BBa_K3187044 )</a> encodes the fusion protein of coat protein, Strep-Tag and LPETGG. The composite part can therefore be used for the production of P22 Virus-like particles (VLPs), as they consist of coat and scaffold protein.

    The scaffold fusion protein also includes a Strep-Tag for purification, as well as sfGFP. As a result of the fusion of sfGFP with the scaffold protein the VLPs will contain sfGFP as a cargo on the inside.

    The coat fusion protein also includes a Strep-Tag <a href="https://parts.igem.org/Part:BBa_K3187025" target="_blank">(BBa_K3187025 )</a> for purification and a LPETGG tag<a href="https://parts.igem.org/Part:BBa_K3187019" target="_blank">(BBa_K3187019 )</a> . As a result of the fusion of the LEPTGG tag with the coat protein the VLPs will present the tag on the outside. This tag is the recognition sequence of Sortase A7M<a href="https://parts.igem.org/Part:BBa_K3187028" target="_blank">(BBa_K3187028 )</a> which catalyzes a covalent bond of the LPETGG and a poly G tag. On this basis the VLPs can be modified on the outside using SortaseA7M after the assembly of the coat and scaffold proteins.

    The expression levels of both sites were characterized using the <a href="https://parts.igem.org/Part:BBa_K3187028" target="_blank">BBa_K3187011</a>.





    Sequence and Features


    Assembly Compatibility:
    • 10
      INCOMPATIBLE WITH RFC[10]
      Illegal EcoRI site found at 76
      Illegal XbaI site found at 2900
      Illegal PstI site found at 1334
    • 12
      INCOMPATIBLE WITH RFC[12]
      Illegal EcoRI site found at 76
      Illegal NheI site found at 1339
      Illegal PstI site found at 1334
    • 21
      INCOMPATIBLE WITH RFC[21]
      Illegal EcoRI site found at 76
      Illegal BamHI site found at 796
      Illegal XhoI site found at 1499
    • 23
      INCOMPATIBLE WITH RFC[23]
      Illegal EcoRI site found at 76
      Illegal XbaI site found at 2900
      Illegal PstI site found at 1334
    • 25
      INCOMPATIBLE WITH RFC[25]
      Illegal EcoRI site found at 76
      Illegal XbaI site found at 2900
      Illegal PstI site found at 1334
      Illegal NgoMIV site found at 1185
    • 1000
      INCOMPATIBLE WITH RFC[1000]
      Illegal SapI.rc site found at 94